The present invention relates to novel five-membered heterocyclic amidines, to their preparation and to their use as competitive inhibitors of trypsin-like serine proteases, especially thrombin and kininogenases such as kallikrein. The invention also relates to pharmaceutical compositions which contain the compounds as active ingredients, and to the use of the compounds as thrombin inhibitors, anticoagulants and antiinflammatory agents.
Thrombin belongs to the group of serine proteases and plays a central part in the blood coagulation cascade as terminal enzyme. Both the intrinsic and the extrinsic coagulation cascade lead via a plurality of amplifying stages to the production of thrombin from prothrombin. Thrombin-catalyzed cleavage of fibrinogen to fibrin then initiates blood coagulation and aggregation of platelets which, in turn, due to the binding of platelet factor 3 and coagulation factor XIII, and a large number of highly active mediators, enhance thrombin formation.
The formation and action of thrombin are central events in the development both of white, arterial and of red, venous thrombi and are therefore potentially effective points of attack for drugs. Thrombin inhibitors are, by contrast with heparin, able independently of cofactors completely to inhibit simultaneously the effects of free thrombin and of that bound to platelets. They are able to prevent in the acute phase thromboembolic events after percutaneous transluminal coronary angioplasty (PTCA) and lysis, and to act as anticoagulants in extracorporeal circulation (heart-lung machine, hemodialysis). They can also be used generally for the prophylaxis of thrombosis, for example after surgical operations.
It is known that synthetic arginine derivatives influence the enzymatic activity of thrombin by interacting with the active serine residue of the protease thrombin. Peptides based on Phe-Pro-Arg in which the N-terminal amino acid is in the D form have-proven particularly beneficial. D-Phe-Pro-Arg isopropyl ester is described as a competitive thrombin inhibitor (C. Mattson et al., Folia Haematol, 109 (1983) 43-51).
Derivatization of the arginine at the C terminus to the aldehyde leads to an enhancement of the inhibitory effect. Thus, a large number of arginals able to bind the hydroxyl group of the xe2x80x9cactivexe2x80x9d serine in a hemiacetal have been described (EP 185390, 479489, 526877, 542525; WO 93/15756, 93/18060.
The thrombin-inhibitory activity of peptide ketones, fluorinated alkyl ketones and of keto esters, boric acid derivatives, phosphoric esters and xcex1-keto carboxamides can likewise be explained by this serine interaction (EP 118280, 195212, 362002, 364344, 410411, 471651, 589741, 293881, 503203, 504064, 530167; WO 92/07869, 94/08941).
The peptide 4-amidinophenylglycinephosphonate diphenyl esters described by J. Oleksyszyn et al. in J. Med. Chem. 37 (1994) 226-231 are irreversible thrombin inhibitors with inadequate selectivity in respect of other serine proteases.
DE 3 108 810, WO 93/11152 and EP 601 459 describe agmatine and hence arginine derivatives which are unable to interact with the active serine in serine proteases.
WO 94/29336, EP 0 601 459 and WO 95/23609 represent a further development in which the agmatine is replaced by an arylamidine residue.
EP 0 672 658 describes not only thrombin inhibitors which have attached to them an agmatine or benzamidine residue, but also a thrombin inhibitor having an amidinothiophene (Example 65).
Kininogenases are serine proteases which liberate vasoactive peptides, called kinins (bradykinin, kallidin and Met-Lys-bradykinin), from kininogens. Kininogens are multifunctional proteins which occur in coagulation and inflammation cascade reactions. As inhibitors, they protect cells from damage by cysteine proteases (Mxc3xcller Esterl, FEBS Lett. 182 (1985) 310-314). Important kininogenases are plasma kallikrein, tissue kallikrein and mast cell tryptase.
Kinins like bradykinin and kallidin are vasoactive peptides which influence a large number of biological processes. They play an essential part in inflammatory processes. By increasing vascular permeability, they lead to hypotension and edema. Furthermore, they are very potent pain-producing substances produced naturally in the body and have great importance as cellular mediators in the pathophysiology of asthma, of allergic rhinitis and of arthritis (K. D. Bhoola, C. D. Figueroa, K. Worthy, Pharmacological Reviews 44 (1) (1992) 1-80).
Irrespective of the mechanisms underlying inflammatory processes, fluid containing all the protein systems in the circulating blood escapes from blood vessels. This means that escape of plasma fluid from vessels is involved in diseases such as asthma, rhinitis and inflammatory internal diseases. Moreover, mast cell tryptase is released particularly in allergic processes (Salomonsson et al., Am. Rev. Respir. Dis. 146 (1992) 1535-1542).
The arginine chloromethyl ketones H-(D)-Pro-Phe-Arg-CH2Cl and H-(D)-Phe-Phe-Arg-CH2-Cl have been described by Kettner and Shaw as plasma kallikrein inhibitors (Biochem. 17 (1978) 4778-4784 and Meth. Enzym. 80 (1981) 826-842).
Various synthetic derivatives of benzamidines and benzylamines have proven to be inhibitors of plasma kallikrein, with the benzamidines having a considerably stronger inhibitory effect (F. Markward, S. Drawert, P. Walsmann, Biochemical Pharmacology 23 (1974) 2247-2256).
PKSI-527, the hydrochloride of N-(trans-4-aminomethylcyclohexylcarbonyl)-L-phenylalanine 4-carboxymethylanilide, is also an effective inhibitor of this kininogenase (Wanaka, Ohamoto et al., Thromb. Res., 57 (6) (1990) 889-895).
The invention relates to compounds of the formula I
Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94Fxe2x80x83xe2x80x83I
in which A, B, D, E and F have the following meanings:
A: 
xe2x80x83where
m is 0, 1 or 2,
n is 0, 1 or 2,
R1 is HOOCxe2x80x94, C1-6-alkyl-OOCxe2x80x94, aryl-C0-4-alkyl-OOC or xe2x80x94OH,
R2 is Hxe2x80x94, C1-4-alkyl- or R1xe2x80x94(CH2)mxe2x80x94 and
R3 is Hxe2x80x94 or C1-4-alkyl-,
B: 
xe2x80x83where
R4 is Hxe2x80x94, C1-4-alkyl- or R1xe2x80x94(CH2)mxe2x80x94 (where R1 and m have the abovementioned meanings),
p is 0 or 1,
R5 is Hxe2x80x94 or C1-4-alkyl-,
R6 is Hxe2x80x94, C1-8-alkyl-, 2-thienyl-, 3-thienyl-, 3-indolyl-, 4-imidazolyl-, 2-pyridyl-, 3-pyridyl-, 4-pyridyl-, phenyl- which may carry up to three identical or different radicals from the group of C1-4-alkyl-, CF3xe2x80x94, C1-4-alkoxy-, HOxe2x80x94, BnOxe2x80x94, Fxe2x80x94 or Clxe2x80x94, or C3-8-cycloalkyl- which may carry up to four identical or different C1-4-alkyl radicals and/or where one or two Cxe2x80x94C single bonds in the ring can be replaced by a Cxe2x95x90C double bond and/or a phenyl ring can be fused on, C7-C12-bicycloalkyl- or C10-tricycloalkyl- or
R4 and R6 together are an ethylene or propylene group,
R7 is H, C1-8-alkyl-, phenyl- which may carry up to three identical or different radicals from the group of C1-4-alkyl-, CF3xe2x80x94, C1-4-alkoxy-, Fxe2x80x94 or Clxe2x80x94, or C3-8-cycloalkyl- which may carry up to four identical or different C1-4-alkyl radicals, and
R8 is H or C1-4-alkyl,
D: 
where R20 is H, C1-4-alkyl, Bn or BnO(CO)xe2x80x94 and where the following applies:
if D is II, III or XI, then E has the following meaning: 
where
a) in the event that X=S, O, NH or NR12,
Y is xe2x80x94CR13xe2x95x90, xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR14xe2x95x90
or
Y is xe2x80x94CR13xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
b) in the event that X=NR12,
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
c) in the event that X=S, O or NH,
Y is xe2x80x94CR15xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR15xe2x95x90
or
d) in the event that X=xe2x80x94NR12xe2x80x94,
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90, xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
and
R9 is Hxe2x80x94 or C1-3-alkyl-,
R10 is Hxe2x80x94 or C1-4-alkyl-,
R11 is Hxe2x80x94 or C1-4-alkyl-,
R12 is CH3xe2x80x94 or C2H5xe2x80x94,
R13 is Clxe2x80x94, CF3xe2x80x94 or C1-4alkyl-,
R14 is Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
R15 is CF3xe2x80x94 or C1-4-alkyl-,
R16 is Hxe2x80x94, CF3xe2x80x94 or C1-4-alkyl- and
R20 is as above,
or, if D is IV, VI, VII, VIII, IX or X, then E has the following meaning: 
where
X is O, S or xe2x80x94NR17xe2x80x94
and
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90 or xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR18xe2x95x90 and
Z is xe2x80x94CR19xe2x95x90
and
R9, R10, R11, R16 and R20 are as above,
R17 is H, CH3xe2x80x94 or C2H5xe2x80x94,
R18 is Hxe2x80x94, Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
R19 is Hxe2x80x94, Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
or
if D is II, III, IV, VI, VII, VIII, IX, X or XI, E has the following meanings: 
where
a) in the event that X=S,
Y is xe2x80x94CR18xe2x95x90 and
Z is xe2x80x94CR19xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
b) in the event that X=O or xe2x80x94NR12xe2x80x94,
Y is xe2x80x94Nxe2x95x90, xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90, xe2x80x94CR18xe2x95x90
and
R9, R10, R11, R12, R16, R18, R19 and R20 have the abovementioned meanings,
F: 
and their salts with physiologically acceptable acids.
The amino acid derivatives represented by B preferably have the (D) configuration; azetidinecarboxylic acid, proline and pipecolic acid in D preferably have the (L) configuration.
Preferred compounds of the formula I are those where A to E have the following meanings:
A:
HOOCxe2x80x94(CH2)txe2x80x94 (t=1, 2 or 3), (HOOCxe2x80x94CH2)2xe2x80x94CHxe2x80x94,
HOOCxe2x80x94CH2xe2x80x94CH(COOH)xe2x80x94, HOOCxe2x80x94CH(C1-4-alkyl)-,
HOOCxe2x80x94C(C1-4-alkyl)2xe2x80x94, C1-6-alkyl-OOCxe2x80x94(CH2)txe2x80x94,
B is 
p is 0 or 1,
R4 is Hxe2x80x94, C1-4-alkyl- or HOOCxe2x80x94(CH2)mxe2x80x94 (m=1, 2 or 3),
R5 is Hxe2x80x94, methyl-
R6 is Hxe2x80x94, C1-8-alkyl-, 2-thienyl-, 3-thienyl-, 3-indolyl-, 4-imidazolyl-, 2-pyridyl-, 3-pyridyl-, 4-pyridyl-, phenyl- which may carry up to three identical or different radicals from the group of CH3xe2x80x94, CF3xe2x80x94, CH3xe2x80x94Oxe2x80x94, HOxe2x80x94, BnOxe2x80x94, Fxe2x80x94 or Clxe2x80x94, or C3-8-cycloalkyl, which may carry up to four methyl radicals, bicyclo[2.2.2]octyl-, bicyclo[2.2.1]heptyl-, adamantyl-, indanyl-, decalinyl-,
R7 is H, C1-8-alkyl-, phenyl-, which may carry up to three identical or different radicals from the group of CH3xe2x80x94, CF3xe2x80x94, CH3Oxe2x80x94, Fxe2x80x94 or Clxe2x80x94, or C3-8-cycloalkyl- which may carry up to four methyl radicals,
R8 is H, C1-4-alkyl,
D: 
where R20 is H, CH3, Bn oder BnO(CO)xe2x80x94 and
where the following applies:
if D is II, III or XI, then E has the meaning: 
where
a) in the event that X=S, O or NR17,
Y is xe2x80x94CR13xe2x95x90 or xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR14xe2x95x90
or
Y is xe2x80x94CR13xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
b) in the event that X=NR12,
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94Hxe2x95x90,
or
c) in the event that X=S, O or NH,
Y is xe2x80x94CR15xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR15xe2x95x90
or
d) in the event that X=NR12,
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90, xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
and
R12 is CH3xe2x80x94 or C2H5xe2x80x94,
R13 is Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
R14 is Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
R15 is CF3xe2x80x94 or C1-4alkyl-,
R16 is Hxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
R17 is H, CH3xe2x80x94 or C2H5xe2x80x94
R20 is as above, or
if D is IV, VI, VII, VIII, IX or X, then E has the meaning: 
where
X is O, S or xe2x80x94NR17xe2x80x94 and where
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90 or xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR18xe2x95x90 and
Z is xe2x80x94CR19xe2x95x90
and
R16, R17, R20 have the abovementioned meanings,
R18 is Hxe2x80x94, Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl- and
R19 is Hxe2x80x94, Clxe2x80x94, CF3xe2x80x94 or C1-4-alkyl-,
or
if D is II, III, IV, VI, VII, VIII, IX, X or XI, then E has the meanings: 
where
a) in the event that X=S,
Y is xe2x80x94CR18xe2x95x90 and
Z is xe2x80x94CR19xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
b) in the event that X=O or xe2x80x94NR12xe2x80x94,
Y is xe2x80x94Nxe2x95x90 or xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90 or xe2x80x94CR18xe2x95x90
and R12, R16, R18, R19 and R20 have the abovementioned meanings,
F: 
and their salts with physiologically acceptable acids.
The amino acid derivatives represented by B preferably have the (D) configuration; azetidinecarboxylic acid, proline and pipecolic acid in D preferably have the (L) configuration.
Especially preferred compounds of the formula I are those where A, B, D, E and F have the following meanings:
A: HOOCxe2x80x94CH2, HOOCxe2x80x94CH2xe2x80x94CH2, HOOCxe2x80x94CH(CH3), HOOCxe2x80x94CH(C2H5) 
p is 0 or 1,
R4 is Hxe2x80x94, CH3xe2x80x94
R5 is Hxe2x80x94, CH3xe2x80x94,
R6 is C1-8-alkyl-, C5-8-cycloalkyl- which may carry up to four methyl radicals, 2-thienyl-, 3-indolyl-, 4-imidazolyl-, 2-pyridyl-, 3-pyridyl-, 4-pyridyl, phenyl- which may carry up to three identical or different radicals from the group of CH3xe2x80x94, CF3xe2x80x94, CH3Oxe2x80x94, HOxe2x80x94, BnOxe2x80x94, Fxe2x80x94 or Clxe2x80x94, bicyclo[2.2.2]octyl, bicyclo[2.2.1]heptyl, adamantyl, indanyl, decalinyl, with particular emphasis on cyclopentyl, cyclohexyl and cycloheptyl,
R7 is H, CH3xe2x80x94,
R8 is H, CH3xe2x80x94,
D: 
where R20 is H, BnO(CO)xe2x80x94 and
where the following applies:
if D is II, III or XI, then E has the meaning 
where
X is xe2x80x94Sxe2x80x94 and where
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR13xe2x95x90
or
Y is xe2x80x94CR13xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
Y is xe2x80x94CR15xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR15xe2x95x90
and
R13 is Clxe2x80x94, CF3xe2x80x94 or CH3xe2x80x94
R15 is CF3xe2x80x94 or CH3xe2x80x94 and
R20 is as above,
or
if D is IV, VI, VII, VIII, IX or X, then E has the meaning: 
where
X is S and where
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR13xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR13xe2x95x90
or
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CHxe2x95x90
and
R13, R20 have the abovementioned meanings and
R16 is Hxe2x80x94, CF3xe2x80x94 or CH3xe2x80x94
or
if D is II, III, IV, VI, VII, VIII, IX, X or XI, then E has the meanings: 
where either
a) in the event that X=S,
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR18xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CR18xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
b) in the event that X=O or NCH3 
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
c) in the event that X=xe2x80x94NR12xe2x80x94
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR18xe2x95x90
and
R12 is CH3xe2x80x94 or C2H5xe2x80x94 and
R18 is H, Clxe2x80x94, CF3xe2x80x94 or CH3xe2x80x94, and
R16, R20 have the abovementioned meanings
F: 
and their salts with physiologically acceptable acids.
The amino acid derivatives represented by B preferably have the (D) configuration; azetidinecarboxylic acid, proline and pipecolic acid in D preferably have the (L) configuration.
Very especially preferred compounds of the formula I are those where A, B, D, E and F have the following meanings:
A: HOOCxe2x80x94CH2, HOOCxe2x80x94CH2xe2x80x94CH2, HOOCxe2x80x94CH(CH3), HOOCxe2x80x94CH(C2H5)
B: 
p is 0 or 1,
R4 is Hxe2x80x94,
R5 is Hxe2x80x94,
R6 is C1-8-alkyl-, 2-thienyl-, 3-indolyl-, 4-imidazolyl-, 2-pyridyl-, 3-pyridyl-, 4-pyridyl-, C5-8-cycloalkyl- which may carry up to four methyl radicals, phenyl- which may carry up to three identical or different radicals from the group of CH3xe2x80x94, CF3xe2x80x94, CH3Oxe2x80x94, HOxe2x80x94, BnOxe2x80x94, Fxe2x80x94 or Clxe2x80x94, bicyclo[2.2.2]octyl, bicyclo[2.2.1]heptyl, adamantyl, indanyl, decalinyl, with particular emphasis on cyclopentyl-, cyclohexyl- and cycloheptyl-,
R7 is H,
R8 is H,
D: 
where the following applies:
if D is II, III or XI, then E has the meaning 
where
X is S and
Y is xe2x80x94CR13xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR13xe2x95x90
or
Y is xe2x80x94CR15xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR15xe2x95x90
and
R13 is Clxe2x80x94, CF3xe2x80x94 or CH3xe2x80x94 and
R15 is CF3xe2x80x94 or CH3xe2x80x94,
or
if D is IV, VI, VII, VIII, IX or X, then E has the meaning 
where
X is S and
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR13xe2x95x90
or
Y is xe2x80x94CR13xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CHxe2x95x90
and
R13 has the abovementioned meaning and
R16 is H, CF3xe2x80x94 or CH3xe2x80x94, or
if D is II, III, IV, VI, VII, VIII, IX, X or XI, then E has the meanings 
where
a) in the event that X=S,
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR18xe2x95x90
or
Y is xe2x80x94CR18xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94Nxe2x95x90
or
b) in the event that X=O or NCH3 
Y is xe2x80x94CHxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90
or
Y is xe2x80x94CR16xe2x95x90 and
Z is xe2x80x94CHxe2x95x90
or
c) in the event that X=NCH3 
Y is xe2x80x94Nxe2x95x90 and
Z is xe2x80x94CR16xe2x95x90
and
R16 has the abovementioned meaning and
R18 is H, Clxe2x80x94 CF3xe2x80x94 or CH3xe2x80x94,
F: 
and their salts with physiologically acceptable acids.
The amino acid derivatives represented by B preferably have the (D) configuration; azetidinecarboxylic acid, proline and pipecolic acid in D preferably have the (L) configuration.
With the exception of the compounds mentioned in the Examples, the following substances must be very especially emphasized:
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-5-(2-am-3-CF3)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(2-am-3-CF3)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-5-(2-am-4-Me)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-5-(2-am-4-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-5-(2-am-4-CF3)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(2-am-4-Me)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(2-am-4-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(2-am-4-CF3)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(2-am-3,4-Me2)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CR2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(5-am)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(5-am-3-Me)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(5-am-3-Me)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(5-am-3-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(5-am-3-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(5-am-4-Me)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(5-am-4-Me)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(5-am-4-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(5-am-4-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Chea-Pro-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CH2-(D)-Cpa-Pro-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am-5-Me)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am-5-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am-5-CF3)-thioph
HOOCxe2x80x94CH2-CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CH2-CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(4-am)-thioph
HOOCxe2x80x94CH2-(D)-Chea-Pro-NHxe2x80x94CH2-4-(2-am)-thioph
HOOCxe2x80x94CH2-(D)-Cpa-Pro-NHxe2x80x94CH2-4-(2-am)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-4-(2-am)-thioph
HOOCxe2x80x94CH2-(D)-Cheg-Pro-NHxe2x80x94CH2-4-(2-am)-thioph
HOOCxe2x80x94CH2-(D)-Cpg-Pro-NHxe2x80x94CH2-4-(2-am)-thioph
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-2-(4-am)-thiaz
MeOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Cpg-Pro-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Chea-Pro-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Cheg-Pro-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am)-thiaz
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(4-am-5-Me)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am-5-Me)-thiaz
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(4-am-5-CF3)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am-5-CF3)-thiaz
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(5-am-4-Me)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(5-am-4-Me)-thiaz
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(5-am-4-CF3)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(5-am-4-CF3)-thiaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(3-am)-isox
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-2-(4-am)-oxaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am)-oxaz
HOOCxe2x80x94CH2-CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-CH2-(D)-Cha-Pro-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-(D)-Chea-Pro-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-CH2-(D)-Cha-Pic-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-CH2-(D)-Chg-Pic-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-5-(3-am)-fur
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-2-(4-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-2-(4-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(4-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(4-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-2-(5-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-2-(5-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-2-(5-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-2-(5-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-2-(5-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-4-(2-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-4-(2-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-4-(2-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-4-(2-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-4-(2-am-1-Me)-pyrr
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-5-(3-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-5-(3-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(3-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-5-(3-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-5-(3-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-3-(5-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-3-(5-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-3-(5-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-3-(5-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Cha-Pic-NHxe2x80x94CH2-3-(5-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Pic-NHxe2x80x94CH2-3-(5-am-1-Me)-pyraz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(3-am)-oxadiaz
tBuOOCxe2x80x94H2-N-BOC-(D)-Chg-Pro-NHxe2x80x94CH2-2-(4-am)-oxaz
HOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(3-am-4-Cl)-thioph
EtOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-(3am-4-Cl)-thioph
HOOCxe2x80x94CH2-(D)-Chg-Pro-NH-H2-5-(3-am-4-Me)-thioph
EtOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-5-3-am-4-Me)-thioph
tBuOOCxe2x80x94CH2-(D)-Cha-Pro-NHxe2x80x94CH2-4-(2-ham)-thioph
tBuOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-4-(2-ham)-thioph
tBuOOCxe2x80x94H2-(D)-Chg-Aze-NHxe2x80x94CH2-4-(2-ham)-thioph
tBuOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-4-(2-ham)-thioph
tBuOOCCH2-(D)-Cha-Pro-NHxe2x80x94CH2-4-(2-ham)-thiaz
tBuOOCxe2x80x94CH2-(D)-Chg-Pro-NHxe2x80x94CH2-4-(2-ham)-thiaz
tBuOOCxe2x80x94CH2-(D)-Chg-Aze-NHxe2x80x94CH2-4-(2-ham)-thiaz
tBuOOCxe2x80x94CH2-(D)-Cha-Aze-NHxe2x80x94CH2-4-(2-ham)-thiaz
In the description and the claims, the following definitions apply to the individual substituents:
The term xe2x80x9ccycloalkylxe2x80x9d, on its own or as part of another substituent comprises saturated or cyclic hydrocarbon groups which contain the given number of carbon atoms. C3-8-cycloalkyl refers to saturated alicyclic rings having 3 to 8 C atoms such as, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 4-methylcyclohexyl, cycloheptyl or cyclooctyl.
The term xe2x80x9calkylxe2x80x9d on its own or as part of another substituent denotes a linear or branched alkyl chain radical of the length indicated in each case. Thus, C1-4-alkyl is, for example, methyl, ethyl, 1-propyl, 2-propyl, 2-methyl-2-propyl, 2-methyl-1-propyl, 1-butyl, 2-butyl, C1-6-alkyl, for example C1-4-alkyl, pentyl, 1-pentyl, 2-pentyl, 3-pentyl, 1-hexyl, 2-hexyl, 3-hexyl, 4-methyl-1-pentyl or 3,3-dimethylbutyl. In addition to the radicals given for C1-4-alkyl, C1-8-alkyl denotes, for example, C1-6-alkyl, heptyl or octyl.
The term xe2x80x9calkoxyxe2x80x9d on its own or as part of another substituent denotes a linear or branched alkyl chain radical which has the length indicated in each case and which is bonded to the respective basic compound via an oxygen atom. Thus C1-4-alkoxy denotes, for example, methoxy, ethoxy, 1-propoxy, 2-propoxy, 2-methyl-2-propoxy, 2-methyl-1-propoxy, 1-butoxy, 2-butoxy.
The invention furthermore relates to compounds which contain the structural element 
where D and E have the abovementioned meanings and where a hydrogen atom, a protective group, an unsubstituted or substituted natural or unnatural amino acid, an unsubstituted or substituted carboxylic acid or an unsubstituted or substituted alkyl radical is located on the nitrogen atom of building block D. The structural fragment is valuable as a component of serine protease inhibitors and, in particular, of thrombin and kallikrein inhibitors.
The invention also relates to compounds which contain the structural element 
where E has the abovementioned meaning and where a hydrogen atom, a protective group, an unsubstituted or substituted natural or unnatural amino acid, an unsubstituted or substituted carboxylic acid or an unsubstituted or substituted alkyl radical is located on the nitrogen atom of NR9.
Finally, the invention also relates to compounds which have one of the following structural elements: 
where Q is CH3 or Cl; T is NCH3, O or S; and W is NCH3 or S.
The invention furthermore relates to the intermediates of the formulae Va and Vb
Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CNxe2x80x83xe2x80x83Va,
Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CSNH2xe2x80x83xe2x80x83Vb,
where A, B, D and E have the abovementioned meanings.
The novel intermediates are used to prepare the compounds I and are valuable building blocks for synthesizing serine protease inhibitors.
The compounds of the formula I can exist as such or in the form of their salts with physiologically acceptable acids. Examples of such acids are: hydrochloric acid, citric acid, tartaric acid, lactic acid, phosphoric acid, methanesulfonic acid, acetic acid, formic acid, maleic acid, fumaric acid, succinic acid, hydroxysuccinic acid, sulfuric acid, glutaric acid, aspartic acid, pyruvic acid, benzoic acid, glucuronic acid, oxalic acid, ascorbic acid and acetylglycine.
If, in the compounds of the formula I, R1 equals C1-6-alkyl-OOC, aryl-C0-4-alkyl-OOC and/or F equals hydroxyamidine, these compounds may act in vivo as prodrugs from which the corresponding carboxylic acids R1xe2x95x90HOOCxe2x80x94 or the corresponding amidines Fxe2x95x90xe2x80x94C(xe2x95x90NH)-NH2 are formed enzymatically.
Prodrugs of the compounds of the formula I are to be understood as meaning those compounds which are metabolized in vivo to give the pharmacologically active compounds of the formula I. This can be effected, for example, by the first-pass metabolism in the liver.
The novel compounds of the formula I are competitive inhibitors of trypsin-like serine proteases, especially of thrombin, and also of kininogenases such as kallikrein. They can be employed for the following indications:
diseases whose pathogenetic mechanism derives directly or indirectly from the proteolytic effect of thrombin,
diseases whose pathogenetic mechanism derives from thrombin-dependent activation of receptors and signal transductions,
diseases associated with stimulation [eg. by PAI-1, PDGF (platelet derived growth factor), P-selectin, ICAM-1, tissue factor] or inhibition (eg. NO synthesis in smooth muscle cells) of the expression of genes in body cells,
diseases deriving from the mitogenic effect of thrombin,
diseases deriving from a thrombin-dependent change in the contractility and permeability of epithelial cells (eg. vascular endothelial cells),
thrombin-dependent thromboembolic events such as deep vein thrombosis, pulmonary embolism, myocardial or cerebral infarct, atrial fibrillation, bypass occlusion,
disseminated intravascular coagulation (DIC),
reocclusion and for reducing the reperfusion time on comedication with thrombolytics such as streptokinase, urokinase, prourokinase, t-PA, APSAC, plasminogen activators from the salivary glands of animals, and the recombinant and mutated forms of all these substances,
the occurrence of early reocclusion and late restenosis after PTCA,
the thrombin-dependent proliferation of smooth muscle cells,
the accumulation of active thrombin in the CNS (eg. in Alzheimer""s disease),
tumor growth and to prevent adhesion and metastasis of tumor cells.
The novel compounds can be used in particular for the therapy and prophylaxis of thrombin-dependent thromboembolic events such as deep vein thromboses, pulmonary embolisms, myocardial or cerebral infarcts and unstable angina, also for the therapy of disseminated intravascular coagulation (DIC). They are furthermore suitable for combination therapy with thrombolytics such as streptokinase, urokinase, prourokinase, t-PA, APSAC and other plasminogen activators to shorten the reperfusion time and extend the reocclusion time.
Further preferred areas of use are to prevent thrombin-dependent early reocclusion and late restenosis after percutaneous transluminal coronary angioplasty, to prevent thrombin-induced proliferation of smooth muscle cells, to prevent accumulation of active thrombin in the CNS (eg. in Alzheimer""s disease), to control tumors and to prevent mechanisms which lead to adhesion and metastasis of tumor cells.
The novel compounds can also be used for coating artificial surfaces such as hemodialysis membranes and the tubing systems and lines necessary therefor, and for coating oxygenators in extravascular circulation, stents and heart valves.
The novel compounds can furthermore be employed for diseases whose pathogenetic mechanism derives directly or indirectly from the proteolytic effect of kininogenases, especially kallikrein, eg. in inflammatory diseases such as asthma, pancreatitis, rhinitis, arthritis, urticaria and other internal inflammatory diseases.
The compounds according to the invention can be administered in a conventional way orally or parenterally (subcutaneously, intravenously, intramuscularly, interperitoneally, rectally). Administration can also take place with vapors or sprays through the nasopharyngeal space.
The dosage depends on the age, condition and weight of the patient and on the mode of administration. As a rule, the daily dose of active substance per person is about 10-2000 mg on oral administration and about 1-200 mg on parenteral administration. This dose can be given in 2 to 4 single doses or once a day as depot form.
The novel compounds can be used in conventional solid or liquid pharmaceutical forms, eg. as uncoated or (film-)coated tablets, capsules, powders, granules, sugar-coated tablets, suppositories, solutions, ointments, creams or sprays. These are produced in a conventional manner. The active substances can for this purpose be mixed with conventional pharmaceutical auxiliaries such as tablet binders, bulking agents, preservatives, tablet disintegrants, flow regulators, plasticizers, wetting agents, dispersants, emulsifiers, solvents, release-slowing agents, antioxidants and/or propellant gases (cf. H. Sucker et al.: Pharmazeutische Technologie, Thieme-Verlag, Stuttgart, 1978). The administration forms obtained in this way normally contain from 0.1 to 99% by weight of active substance.
The compounds of the formula I can be prepared as shown in Schemes I-III.
Building blocks A, B, D and E are preferably assembled separately beforehand and employed in suitably protected form (see Scheme I-III). 
Scheme I describes the linear assemblage of the molecule I by coupling the amine Hxe2x80x94Exe2x80x94CN to the N-protected amino acid Pxe2x80x94Dxe2x80x94OH to give Pxe2x80x94Dxe2x80x94Exe2x80x94CN, eliminating the N-terminal protective group to give Hxe2x80x94Dxe2x80x94Exe2x80x94CN, coupling to the N-protected amino acid Pxe2x80x94Bxe2x80x94OH to give Pxe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN, eliminating the protective group P to give Hxe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN, subsequently alkylating with the unprotected or protected (P)xe2x80x94Axe2x80x94U building block (U=leaving group) or reductively alkylating with (P)xe2x80x94Axe2x80x2xe2x80x94U (U=aldehyde, ketone) or Michael addition with a suitable (P)xe2x80x94Axe2x80x3xe2x80x94Cxe2x95x90Cxe2x80x94 derivative to give (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN. Conversion of the nitrile functionality into the amidine group takes place either by the classical Pinner synthesis (R. Boder, D. G. Neilson, Chem. Rev. 61 (1962) 179) or by a modified Pinner synthesis which proceeds via imino thioester salts as intermediate (H. Vieweg et al., Pharmazie 39 (1984) 226) or directly by the method of A. Eschenmoser Helv. Chimica Acta 69 (1986) 1224. Subsequently the protective groups still present in the molecule are eliminated, preferably by acid hydrolysis.
If building block E is incorporated as Hxe2x80x94Exe2x80x94CONH2 into the synthesis, dehydration of the amide to the.nitrile functionality or conversion to the thioamide functionality takes place on one of the protected intermediates. As an alternative, the building block E may be employed in the synthesis in the form of Hxe2x80x94Exe2x80x94CSNH2. 
Scheme II describes the linear assemblage of the molecule I by alkylation, reductive amination or Michael addition of Hxe2x80x94Bxe2x80x94P onto appropriately suitable unprotected or protected A building blocks to give (P)xe2x80x94Axe2x80x94Bxe2x80x94P, elimination of the C-terminal protective group to give (P)xe2x80x94Axe2x80x94Bxe2x80x94OH, coupling to Hxe2x80x94Dxe2x80x94P to give (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94P, elimination of the C-terminal protective group to give (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94OH, coupling to Hxe2x80x94Exe2x80x94CN to give (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN and reaction of this intermediate to give the final product as in Scheme I.
Where compounds (P)xe2x80x94Axe2x80x94Bxe2x80x94P still have a free NH functionality on B, this must be provided with a suitable protective group before elimination of the C-terminal protective group. The protective groups used in each case must be orthogonal to one another.
As an alternative to the Hxe2x80x94Exe2x80x94CN building block, it is also possible to employ Hxe2x80x94Exe2x80x94CONH2, Hxe2x80x94Exe2x80x94CSNH2, Hxe2x80x94Exe2x80x94C(NH)NH2, Hxe2x80x94Exe2x80x94C(NP)NH2, Hxe2x80x94Exe2x80x94C(NP)NHP, with the coupled intermediate (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CONH2 in the first case being dehydrated to (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN or being converted directly into (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CSNH2, for example by means of Lawesson""s reagent. 
Scheme III describes a very efficient way for preparing compounds I by a convergent synthesis. The appropriately protected building blocks (P)xe2x80x94Axe2x80x94Bxe2x80x94OH and Hxe2x80x94Dxe2x80x94Exe2x80x94CN are coupled together and the resulting intermediate (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN is reacted to give the final product as in Scheme I.
It is also possible to employ Hxe2x80x94Dxe2x80x94Exe2x80x94CONH2 or Hxe2x80x94Dxe2x80x94Exe2x80x94CSNH2 as an alternative to Hxe2x80x94Dxe2x80x94Exe2x80x94CN, with the coupled intermediate (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CONH2 in the first case being dehydrated to (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CN or being converted into (P)xe2x80x94Axe2x80x94Bxe2x80x94Dxe2x80x94Exe2x80x94CSNH2.
The N-terminal protective groups employed are Boc, Cbz or Fmoc, preferably Boc, and the C-terminal protective groups are methyl, tert-butyl and benzyl. If a plurality of protective groups is present in the molecule, they must be orthogonal to one another if they are not to be eliminated simultaneously.
The required coupling reactions and the other reactions for introducing and eliminating protective groups are carried out under standard conditions of peptide chemistry (see M. Bodanszky, A. Bodanszky xe2x80x9cThe Practice of Peptide Synthesisxe2x80x9d, 2nd edition, Springer Verlag Heidelberg, 1994).
Boc protective groups are eliminated using dioxane/HCl or TFA/DCM, and Cbz protective groups are eliminated by hydrogenolysis or with HF. Hydrolysis of ester functionalities takes place with LiOH in an alcoholic solvent or in dioxane/water. TFA or HCl are used to cleave t-butyl esters.
The reactions were checked by TLC, normally using the following mobile phases:
Where separations by column chromatography are mentioned, these were separations on silica gel using the abovementioned mobile phases.
Reversed phase HPLC separations were carried out with aceto-nitrile/water and HOAc buffer.
All reactions were routinely carried out under a nitrogen atmosphere.
The starting compounds can be prepared by the following methods:
Examples of building blocks A employed for the alkylation are tert-butyl xcex1-bromoacetate, tert-butyl xcex2-bromopropionate, tert-butyl xcex1-bromopropionate, tert-butyl xcex3-bromobutyrate, tert-butyl xcex1-bromobutyrate, THP-protected bromoethanol, THP-protected xcex3-bromopropanol, xcex1-bromo-xcex3-butyrolactone, for the reductive amination are dihydroxyacetone, di-tert-butyl acetonedicarboxylate, and for the Michael addition are tert-butyl acrylate, tert-butyl methacrylate, di-tert-butyl fumarate. Those of said tert-butyl esters which cannot be purchased are prepared by methods similar to G. Uray, W. Lindner, Tetrahedron, 44 1988 357-4362.
B building blocks:
A wide variety of possibilities is available in the literature for the general and specific synthesis of amino acids. A review thereof is provided by, inter alia, Houben-Weyl, Volume E16d/Part 1, pages 406 et seq.
Precursors which were frequently employed were benzophenone imine acetic acid ethyl ester, diethyl-acetamidomalonate and ethyl isonitrileacetate.
Various racemic glycine and alanine derivatives were prepared, for example, starting from ethyl isonitrileacetate and an appropriate ketone or aldehyde (see H.-J. Prxc3xa4torius, J. Flossdorf, M.-R. Kula Chem. Ber. 108 (1975) 3079).
The syntheses of cyclooctylglycine, cycloheptylglycine, 2-norbonylglycine [sic], adamantylalanine, xcex3-methylcyclohexylalanine, 4-isopropyl-1-cyclohexylalanine, 4-methyl-1-cyclohexylalanine, 4-methyl-1-cyclohexylglycine, cycloheptylalanine and cyclopentyl-alanine were carried out via the corresponding ethyl 2-formylaminoacrylates (U. Schxc3x6llkopf and R. Meyer, Liebigs Ann. Chem. 1977, 1174 and H.-J. Prxc3xa4torius, J. Flossdorf, M.-R. Kula Chem. Ber. 108 (1985) 3079) starting from ethyl isocyanoacetate with the relevant carbonyl compounds cyclooctanone, cyclo-heptanone, 2-norbornanone, 1-formyladamantane, 1-formyl-1-methylcyclohexane, 1-formyl-4-isopropylcyclohexane, 1-formyl-4-methylcyclohexane and 4-methyl-cyclohexanone, formylcyclohexane and formylcyclopentane by the following general methods:
A solution of 100 m [sic] of ethyl isocyanoacetate in 50 ml of THF was added dropwise to 100 m [sic] of potassium tert-butoxide in 150 ml of THF at 0 to xe2x88x9210xc2x0 C. After 15 min at the same temperature 100 mmol of the appropriate carbonyl compound in 50 ml of THF were added, the reaction mixture was allowed to rise slowly to RT, and the solvent was stripped off in a rotary evaporator. The residue was mixed with 50 ml of water, 100 ml of acetic acid and 100 ml of DCM, and the product was extracted with DCM. The DCM phase was dried over Na2SO4, and the solvent is stripped off in a rotary evaporator. The resulting products were almost pure but could, if necessary, be purified further by column chromatography on silica gel (mobile phases: ether/petroleum ether mixtures).
100 m [sic] of the ethyl 2-formylaminoacrylates were hydrogenated with Pd/C (10%) and hydrogen in 200 ml of glacial acetic acid until the reaction was complete. The catalyst was then filtered off, the acetic acid was stripped off as far as possible in a rotary evaporator, and the residue was refluxed in 200 ml of 50% concentrated hydrochloric acid for 5 h. The hydrochloric acid was stripped off in a rotary evaporator, and the product was dried at 50xc2x0 C. under reduced pressure and then washed several times with ether. The hydrochlorides resulted as pale colored crystals.
25.0 g of cyclooctylglycine hydrochloride were obtained starting from 18.9 g (150 mmol) of cyclooctanone. 36.2 g of cycloheptylglycine hydrochloride were obtained starting from 22.4 g (200 mmol) of cycloheptanone. 26.6 g of 2-norbonylglycine [sic] hydrochloride were obtained starting from 16.5 g (150 mmol) of 2-norbornanone. 26.0 g of adamantylalanine hydrochloride were obtained starting from 19.7 g (120 mmol) of 1-formyladamantane. 16.6 g of y-methylcyclohexylalanine hydrochloride were obtained starting from 12.6 g (100 mmol) of 1-formyl-1-methylcyclohexane. 25.9 g of 4-methylcyclohexylglycine hydrochloride were obtained starting from 16.8 g (150 mmol) of 4-methylcyclohexanone. 18 g of trans-4-methyl-1-cyclohexylalanine hydrochloride were obtained starting from 15 g of trans-1-formyl-4-methylcyclohexane. 10 g of 3,3-dimethyl-1-cyclohexylalanine hydrochloride were obtained starting from 9 g of 3,3-dimethyl-1-formylcyclohexane.
The aldehyde 1-formyl-3,3-dimethylcyclohexane required for the synthesis was prepared by a method based on that of Moskal and Lensen (Rec. Trav. Chim. Pays-Bas 106 (1987) 137-141).
A solution of n-butyllithium in n-hexane (72 ml, 115 mmol) was added dropwise over the course of 10 min to a stirred solution of diethyl isocyanomethylphosphonate (17 ml, 105 mmol) in 280 ml of anhydrous diethy [sic] ether at xe2x88x9260xc2x0 C. The resulting suspension was then stirred at xe2x88x9260xc2x0 C. for 15 min and, over the course of 10 min, a solution of 3,3-dimethylcyclohexanone (13 g, 105 mmol) in 100 ml of anhydrous diethyl ether was added, keeping the temperature below xe2x88x9245xc2x0 C. The reaction mixture was allowed to reach 0xc2x0 C. and, after stirring at this temperature for 90 min, 150-200 ml of 38% strength aqueous hydrochloric acid were cautiously added. The mixture was vigorously stirred at room temperature for 15 h to complete the hydrolysis. The organic phase was separated off and washed with 200 ml each of water, saturated sodium bicarbonate solution and saturated sodium chloride solution. It was dried over magnesium sulfate, filtered and concentrated in a rotary evaporator in order to remove the solvent. The resulting residue was employed without further purification as starting material for synthesizing the amino acid.
Cyclopentylglycine was prepared by hydrolysing N-acetyl-(D,L)-cyclopentylglycine with 6N hydrochloric acid, the former having been prepared as described in the literature by J. T. Hill and F. W. Dunn, J. Org. chem. 30(1965), 1321.
3.4 g (12.2 mmol) of Boc-(D)-xcex1-methyl-Phe-OH were hydrogenated in 100 ml of MeOH in the presence of 250 mg of 5% Rh on Al2O3 under 10 bar with hydrogen at 50xc2x0 C. for 24 h. Filtration and stripping off the solvent resulted in 2.8 g of Boc-(D)-xcex1-methyl-Cha-OH.
1H NMR (DMSO-d6, xcex4 in ppm): 12 (very broad signal, COOH); 1.7-0.8 (25H; 1.35 (s, Boc), 1.30 (s, Me)).
Boc-(3-Ph)-Pro-OH was synthesized by a method similar to that of J. Y. L. Chung et al. (J. Y. L. Chung et al., J.Org.Chem. 55 (1990) 270).
Boc-1-Tetralinylglycine was prepared starting from 1,2-dihydronaphthalene. 1,2-Dihydronaphthalene was initially converted into 1-tetralyl bromide with HBr (similar to J. Med. Chem. 37 (1994) 1586). The bromide was subsequently reacted with diethyl acetamidomalonate and, after hydrolytic cleavage, the resulting xcex1-amino acid was converted into the Boc-protected form under standard conditions. Another possible preparation is described by E. Reimann and D. Voss (E. Reimann, D. Voss, Arch. Pharm. 310 (1977) 102).
Boc-(D,L)-Dpa-OH (1 mmol) was hydrogenated in 12 ml of MeOH together with catalytic amounts of 5% Rh/Al2O3 under 5 bar. Filtration and removal of the solvent under reduced pressure resulted in the product in quantitative yield. Preparation of H-D,L-Chea-OH
4.0 g of cycloheptylmethyl methanesulfonate (19.39 mmol), prepared from cycloheptylmethanol and methanesulfonyl chloride, were refluxed together with 4.9 g of benzophenone imine glycine ethyl ester (18.47 mmol), 8.9 g of dry, finely powdered potassium carbonate (64.65 mmol) and 1 g of tetrabutylammonium bromide (3 mmol) in 50 ml of dry acetonitrile under an inert gas atmosphere for 10 h. The potassium carbonate was then filtered off, the filtrate was evaporated to dryness, and the crude product was hydrolyzed directly with 20 ml of 2N hydrochloric acid in 40 ml of ethanol, stirring at RT for 1.5 h. The reaction solution was diluted and then benzophenone was extracted with ethyl acetate in the acidic range, and subsequently H-D,L-Chea-OEt was extracted with DCM in the alkaline range (pH=9), and the solution was dried over magnesium sulfate and concentrated in a rotary evaporator. Yield 3.7 g{circumflex over (=)}95% of theory.
Boc-(D,L)-(3,4,5-(MeO)3)Phe-OH was prepared by alkylation of benzophenone imine glycine ethyl ester with trimethoxybenzyl chloride, subsequent introduction of the Boc protective group and ester hydrolysis.
H-(D,L)-P,P-Me2Cha-OH was prepared by the method of U. Schxc3x6llkopf, R. Meyer, L. Ann. Chem. (1977) 1174-82.
Said amino acids were converted with di-tert-butyl dicarbonate in water/dioxane by conventional methods into the Boc-protected form in each case and subsequently recrystallized from ethyl acetate/hexane mixtures or purified by column chromatography on silica gel (mobile phases: ethyl acetate/petroleum ether mixtures).
The Boc-protected amino acids were employed as B building blocks as shown in Scheme I.
Said amino acids as B building blocks were also in some cases converted into the corresponding benzyl esters and linked to the appropriately protected A building blocks. In the case of compounds with an Nxe2x80x94H functionality which was still free, this was subsequently protected with a Boc group, the benzyl ester group was removed by hydrogenation, and the building block Axe2x80x94Bxe2x80x94OH was purified by crystallization, salt precipitation or column chromatography. This route is described by way of example for tBuOOCxe2x80x94CH2xe2x80x94(Boc)(D)Cha-OH below.
A suspension of 100 g (481 mmol) of (D)-cyclohexylalanine hydrochloride, 104 g (962 mmol) of benzyl alcohol and 109.7 g (577 mmol) of p-toluenesulfonic acid monohydrate in 2200 ml of toluene was slowly heated to reflux with a water separator. Evolution of hydrogen chloride and dissolving of the suspension to give a clear solution were observed in the temperature range 80-90xc2x0 C. When no further water separated out (about 4 h), 500 ml of toluene were distilled out, the reaction mixture was allowed to cool overnight, and the resulting residue was filtered off and washed twice with 1000 ml of hexane each time. The resulting residue (195 g) was then suspended in 2000 ml of dichloromethane and, after addition of 1000 ml of water, adjusted to pH 9-9.5 by gradual addition of 50% strength sodium hydroxide solution while stirring. The organic phase was separated off, washed twice with 500 ml of water each time, dried over sodium sulfate and filtered to remove desiccant, and concentration of the filtrate resulted in 115 g (94%) of the title product as pale oil.
115 g (440 mmol) of (D)-cyclohexylalanine benzyl ester were dissolved in 2000 ml of acetonitrile and, at room temperature, 607.5 g (4.40 mol) of potassium carbonate and 94.3 g (484 mmol) of tert-butyl bromoacetate were added, and the mixture was stirred at this temperature for 3 days. Carbonate was filtered off, washed with acetonitrile, the mother liquor was concentrated (30xc2x0 C., 20 mbar), the residue was taken up in 1000 ml of methyl tert-butyl ether, and the organic phase was extracted with 5% strength citric acid and saturated sodium bicarbonate solution. The organic phase was dried over sodium sulfate, filtered to remove desiccant and concentrated, and the resulting oil (168 g) was employed directly in the next reaction.
The oil (168 g, 447 mmol) obtained in the previous synthesis was dissolved in 1400 ml of acetonitrile and, after addition of 618 g (4.47 mmol) of potassium carbonate powder and 107.3 g (492 mmol) of di-tert-butyl dicarbonate, stirred at room temperature for 6 days. The potassium carbonate was filtered off with suction, washed with about 1000 ml of acetonitrile, and the filtrate was concentrated. 230 g of the required product were obtained.
115 g of N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylalaine [sic] benzyl ester were dissolved in 1000 ml of pure ethanol and hydrogenated in the presence of 9 g of 10% Pd on active carbon with hydrogen under atmospheric pressure at 25-30xc2x0 C. for 2 h. Filtration and removal of the solvent in a rotary evaporator resulted in 100 g (260 mmol) of a yellow oil which was taken up in 1600 ml of acetone and heated to reflux. The heating bath was removed, and a solution of 27 g (273 mmol) of cyclohexylamine in acetone was quickly added through a dropping funnel. The required salt crystallized out on cooling the reaction mixture to room temperature. The solid was filtered off, washed with 200 ml of acetone and, for final purification, recrystallized once more from acetone. Drying of the residue in a vacuum oven at about 30xc2x0 C. resulted in 70.2 g of the required salt as white powder.
N-Boc-N-(tert-Butyloxycarbonylmethylene)-(D)-cyclohexylglycine cyclohexylammonium salt was prepared analogously from cyclohexylglycine as precursor. The N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cycloheptylglycine and N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclopentylglycine derivatives were prepared from the corresponding cycloheptyl- and cyclopentylglycine compounds.
a) tert-Butyl 3-Bromopropionate
In a countercurrent of nitrogen, 16.64 g (109 mmol) of bromopropionic acid, 150 ml of condensed 2-methylpropene and 2 ml of concentrated sulfuric acid were added at xe2x88x9230xc2x0 C. into an autoclavable glass vessel, the autoclave was sealed tightly and the mixture was stirred for 72 hours at room temperature. For working up, the reaction vessel was again cooled to xe2x88x9230xc2x0 C. and the reaction solution was poured carefully into 200 ml of an ice-cold saturated sodium hydrogen carbonate solution. Excess 2-methylpropene was evaporated with stirring, the residue was extracted three times with in each case 50 ml of dichloromethane, and the combined organic phases were dried over sodium sulfate, filtered to remove desiccant and concentrated under a water pump vacuum. The oily residue was purified by column chromatography (mobile phase n-hexane, later n-hexan/diethyl ether 9:1). This resulted in 18.9 g of the title compound.
b) N-(tert-Butyloxycarbonylethylene)-(D)-cyclohexylalanine Benzyl Ester
49.4 g (189 mmol) of (D)-cyclohexylalanine benzyl ester were dissolved in 250 ml of acetonitrile, the solution was treated with 31.6 g (151 mmol) of tert-butyl bromopropionate at room temperature and the mixture was refluxed for 5 days. The mixture was filtered to remove the precipitate formed and this was washed repeatedly with acetonitrile, the filtrate was concentrated under a water pump vacuum, the residue was taken up in 350 ml of dichloromethane, and the organic phase was extracted with 5% strength citric acid and saturated sodium hydrogen carbonate solution. The organic phase was dried over sodium sulfate, filtered to remove desiccant and concentrated. The oily residue was purified by column chromatography (mobile phase dichloromethane, later dichloromethane/methanol 95:5). This resulted in a slightly impure oil which was employed directly in the next reaction.
c) N-Boc-N-(tert-Butyloxycarbonylethylene)-(D)-cyclohexylalanine Benzyl Ester
The oil obtained in the synthesis above (30 g, max. 70 mmol) was dissolved in 150 ml of acetonitrile, and the solution was treated with 28 ml (160 mmol) of diisopropylethylamine and 19.2 g (88 mmol) of di-tert-butyl dicarbonate and stirred for 3 days at room temperature. The reaction mixture was concentrated under a water pump vacuum in a rotary evaporator, the residue was taken up in n-hexane, the mixture was washed 5 times using in each case 3 ml of a 5% strength citric acid solution, the combined organic phases were dried over sodium sulfate, filtered to remove desiccant and concentrated, and the residue was subjected to separation by column chromatography (mobile phase hexane/ethyl acetate 95:5). The results were 32.66 g (64 mmol) of the required product.
d) N-Boc-N-(tert-Butyloxycarbonylethylene)-(D)-cyclohexylalanine Cyclohexylammonium Salt 32.66 g (64 mmol) of N-Boc-N-(tert-butyloxycarbonylethylene)-(D)-cyclohexylalanine benzyl ester were dissolved in 325 ml of pure ethanol and hydrogenated with hydrogen for 14 hours under atmospheric pressure at 25-30xc2x0 C. in the presence of 3 g of 10% pure Pd on active charcoal. Filtration of the solution through Celite(copyright), washing of the latter with ethanol and removal of the solvent in a rotary evaporator resulted in 26.7 g of a yellow oil which was taken up in acetone and heated to reflux. The heating bath was removed, and a solution of 7 g (70 mmol) of cyclohexylamine in acetone was quickly added through a dropping funnel. The required salt crystallized out on cooling the reaction mixture to room temperature. The solid was filtered off, washed with 25 ml of acetone and, for final purification, recrystallized once more from acetone. Drying of the residue in a vacuum oven at 30xc2x0 C. resulted in 26.6 g (54 mmol) of the required salt as white powder.
N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylalanyl-3,4-dehydroproline:
a) N-Boc-Pyr-OH (5 g, 23.45 mmol) was dissolved in MeOH (50 ml), and HCl in dioxane (4N, 30 ml) was added. The mixture was subsequently heated under reflux for 12 hours. Removal of the solvent in a rotary evaporator resulted in H-Pyr-OMe-hydrochloride as product. Yield: 3.84 g (100%).
b) N-(t-Bu02Cxe2x80x94CH2)-N-Boc-(D)-Cha-OH (8 g, 20.75 mmol) was dissolved in dichloromethane (75 ml), and ethyldiisopropylamine (15.5 ml, 89.24 mmol) were added at xe2x88x9210xc2x0 C. After the mixture had been stirred for 5 minutes at this temperature, a solution of H-Pyr-OMe hydrochloride (3.4 g, 20.75 mmol) in dichloromethane (25 ml) was added dropwise. A solution of propanephosphonic anhydride in ethyl acetate (50% strength, 20 ml, 26.96 mmol) was subsequently added dropwise and stirred for 2 h at xe2x88x9210 to 0xc2x0 C. The batch was diluted with dichloromethane and washed with saturated sodium hydrogen carbonate solution (2xc3x9780 ml), 5% strength citric acid solution (2xc3x9715 ml) and saturated sodium chloride solution (1xc3x9720 ml). The organic phase was dried over sodium sulfate and the solvent was removed in a rotary evaporator. The crude product was purified by means of flash chromatography (silica gel, dichloromethane/methanol 95/5). Yield: 6.2 g (60%).
c) N-(t-BuO2Cxe2x80x94CH2)-N-Boc-(D)-Cha-Pyr-OMe (5.5 g, 11.12 mmol) was dissolved in dioxane (40 ml), aqueous sodium hydroxide solution (IN, 22.2 ml, 22.24 mmol) was added, and the mixture was stirred for 2 hours at room temperature. The dioxane was removed in a rotary evaporator, and the aqueous phase was washed with ethyl acetate and acidified to pH 1-2 with potassium hydrogen sulfate solution (20% strength). The aqueous phase was extracted with dichloromethane and the combined organic phases were dried over sodium sulfate. Yield: 5 g (94%), colorless foam. Recrystallization of n-hexane saturated with water resulted in the corresponding carboxylic acid as colorless crystals (m.p.=158-160xc2x0 C.).
N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylglycyl-3,4-dehydroproline:
This compound was synthesized analogously from N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylglycine and 3,4-dehydroproline methyl ester.
N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylalanylproline:
a) N-(t-BuO2Cxe2x80x94CH2)-N-Boc-(D)-Cha-OH (20 g, 51.88 mmol) was dissolved in dry methylene chloride (100 ml). After cooling to xe2x88x925xc2x0 C., N-ethyldiisopropylamine (90 ml, 518.88 mmol) was added dropwise and stirring was continued for 5 minutes.
H-Pro-OBnxc3x97HCl (12.54 g, 51.88 mmol) was subsequently added at xe2x88x925xc2x0 C. and, after the mixture had been stirred for 5 minutes, 50% strength propanephosphonic anhydride solution in ethyl acetate (45.1 ml, 62.26 mmol), diluted with methylene chloride (45 ml), was added dropwise in the course of 30 minutes. After the mixture had been stirred for 1 hour at 0-5xc2x0 C., it was slowly brought to RT and stirred for 12 hours at RT. The batch was diluted with methylene chloride and washed in succession with saturated sodium hydrogen carbonate solution, 5% strength citric acid solution and saturated sodium chloride solution. After drying over sodium sulfate, the solvent was distilled off in vacuo. Yield: 28.9 g (pale yellow oil, 97%).
b) The product obtained as described in a) (28.5 g, 49.76 mmol) was dissolved in methanol (650 ml), 10% pure Pd on charcoal (1.8 g) was added, and the mixture was hydrogenated at RT and under 1 atmosphere of hydrogen. The catalyst was subsequently removed by filtration through Celite(copyright) and the filtrate was concentrated in vacuo. Yield: 22.2 g (colorless foam, 92%).
N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylglycylproline:
This compound was prepared analogously from N-Boc-N-(tert-butyloxycarbonylmethylene)-(D)-cyclohexylglycine and proline methyl ester.
D building blocks:
The compounds employed as D building blocks, (L)-proline, (L)-pipecolic acid and (L)-azetidinecarboxylic acid, are commercially available, either as free amino acids, as Boc- protected compounds or as the corresponding methyl esters. If (L)-3,4-dehydroproline or (D,L)-4,5-dehydropipecolic acid, or a corresponding, protected derivative, were employed as D building blocks, the compounds prepared were generally hydrogenated in the last step to give the corresponding proline derivatives. (L)-3,4-Dehydroproline (H-Pyr-OH) is commercially available, (D,L) 4,5-dehydropipecolic acid (H-(D,L)-Dep-OH) can be prepared by the methods of A. Burgstahler, C. E. Aiman J. Org. Chem. 25 (1960), 489 or C. Herdeis, W. Engel Arch. pharm 326 (1993), 297.
The E building blocks were synthesized as follows:
5-Aminomethyl-2-cyanothiophene:
This building block was synthesized as described in WO 95/23609.
a) 2-Bromo-4-formylthiophene
36 g (320 mmol) of 3-formylthiophene were dissolved in 600 ml of methylene chloride, the solution was cooled to 5xc2x0 C., 100 g (750 mmol) of aluminum trichloride were added, a little at a time, and the reaction mixture was subsequently refluxed. A solution of 59 g (19 ml, 360 mmol) of bromine in 40 ml of methylene chloride was added dropwise over the course of 45 minutes and the mixture was allowed to after-react under reflux for 4 hours. After cooling, the reaction solution was poured onto 600 g of ice-water and extracted with methylene chloride, and the organic phase was washed with saturated sodium hydrogen carbonate solution, dried over magnesium sulfate and concentrated in a rotary evaporator in vacuo. 64.5 g of crude product resulted, which was purified by means of column chromatography (silica gel, methylene chloride/petroleum ether). A total of 56.5 g of slightly impure product were obtained.
b) 2-Cyano-4-formylthiophene
7.6 g (85 mmol) of copper(I) cyanide were added to a solution of 13.53 g (70.82 mmol) of 2-bromo-4-formylthiophene in 25 ml of DMF and the reaction mixture was refluxed for 3.5 hours, during which process the suspension, which originally was pale green in color, turned into a black solution. After addition of water, the reaction mixture was extracted repeatedly with ethyl acetate, and the organic phases were combined, washed with saturated sodium chloride solution, dried over sodium sulfate and concentrated under slightly reduced pressure. Treatment of the residue (7 g) with ether resulted in 1.6 g of pure product. The mother liquor together with the crude products from other batches was purified by chromatography. (Silica. gel, methylene chloride/petroleum ether 1:1). In total, 56.5 g of 2-bromo-4-formylthiophene were reacted to give 2-cyano-4-formylthiophene, resulting in 12.6 g of pure product (yield 31%).
c) 2-Cyano-4-hydroxymethylthiophene
3.47 g (91.8 mmol) of sodium borohydride were added, a little at a time, to a suspension of 12.6 g (91.8 mmol) of 2-cyano-4-formylthiophene in 200 ml of ethanol and stirred at room temperature for 2 hours, during which process the reaction mixture slowly formed a clear solution. After concentration in vacuo, the residue was taken up in ethyl acetate, washed in succession with saturated sodium chloride solution, 5% strength citric acid and saturated sodium chloride solution, and the organic phase was dried using sodium sulfate and concentrated in vacuo. 11.7 g of almost pure product resulted (yield 91.5%).
d) 4-Bromomethyl-2-cyanothiophene
11.7 g (84.07 mmol) of 2-cyano-4-hydroxymethylthiophene together with 24.1 g (91.87 mmol) of triphenylphosphine were dissolved in 100 ml of-THF at room temperature, and 30.47 g (91.87 mmol) of tetrabromomethane were added, a little at a time, with cooling (ice bath). After stirring for 3 hours at room temperature, the mixture was concentrated in vacuo and purified by chromatography over silica gel (methylene chloride/petroleum ether). 18.8 g of crystalline pale yellow product which still contained petroleum ether resulted.
e) 4-N,N-bis(tert-Butoxycarbonyl)aminomethyl-2-cyanothiophene
18.81 g of 4-bromomethyl-2-cyanothiophene (crude product, maximum 84.07 mmol) were dissolved in 160 ml of THF, the solution was cooled to 5xc2x0 C., and 3.07 g (102.4 mmol) of 80% sodium hydride suspension were added, a little at a time. 22.25 g (102.4 mmol) of di-tert-butyl iminodicarboxylate, dissolved in 160 ml of THF, were subsequently added dropwise at 5xc2x0 C. and the mixture was then stirred overnight at room temperature. Since TLC revealed that the reaction was incomplete, the batch was heated for 4.5 hours at 30-35xc2x0 C.
After cooling to 0-5xc2x0 C., 33 ml of saturated ammonium chloride solution were slowly added dropwise, THF was distilled off in vacuo, the residue was extracted repeatedly with ethyl acetate, and the ethyl acetate phases were washed with saturated sodium chloride solution, dried over sodium sulfate and evaporated in a rotary evaporator. The red viscous residue (34.61 g) was employed in the subsequent reaction as crude product.
f) 4-Aminomethyl-2-cyanothiophene hydrochloride
34.61 g of 4-N,N-bis(tert-butoxycarbonyl)aminomethyl-2-cyano-thiophene (crude product, maximum 84.07 mmol) were dissolved in 600 ml of ethyl acetate, and the solution was cooled to 0-5xc2x0 C., saturated with HCl gas and warmed to room temperature. After 3 hours, the resulting suspension was evaporated in a rotary evaporator, the product was codistilled repeatedly with methylene chloride, and the residue was extracted by stirring with ether and dried in vacuo. 13.85 g of product resulted as a pale powder. Yield over two steps: 94.3%.
a) 4-Cyanothiophene-2-carbaldehyde
49.3 g (258.05 mmol) of 4-bromothiophene-2-carbaldehyde and 27.8 g (310.41 mmol) of copper(I) cyanide were suspended in 130 ml of absolute DMF and the suspension was refluxed for 8 hours. The solvent was evaporated in vacuo in a rotary evaporator at 400C, the residue was suspended in ethyl acetate and the suspension was transferred into a Soxleth apparatus. The residue was extracted overnight, the yellow solution was dried over sodium sulfate and evaporated in vacuo in a rotary evaporator, and the resulting yellow solid was recrystallized frome ether. 25.3 g of product resulted (80% of theory).
b) 4-Cyanothiophene-2-carbaldehyde oxime
11.6 g (84.6 mmol) of 4-cyanothiophene-2-carbaldehyde were dissolved in 140 ml of methanol and 12.3 g (116.1 mmol) of sodium carbonate were added. 6.5 [lacuna] (93.5 mmol) of hydroxylamine hydrochloride were subsequently added at 15xc2x0 C. with cooling, a little at a time, and the mixture was stirred for 2 hours at 10xc2x0 C. After 80 ml of water had been added, the reaction mixture was extracted five times using in each case 50 ml of diethyl ether, the organic phase was dried over sodium sulfate and the solvent was removed in vacuo. 12.5 g of the required product resulted as a yellow crystal powder (96% of theory).
c) 2-Aminomethyl-4-cyanothiophene Hydrochloride
11.22 g (171.64 mmol) of fine zinc dust were carefully added, in several small portions, to a solution of 4.65 g (30.60 mmol) of 4-cyanothiophene-2-carbaldehyde oxime in 50 ml of trifluoroacetic acid, cooled to 0-5xc2x0 C., in such a way that the temperature did not climb above 15xc2x0 C. After stirring for 3 hours at room temperature, excess zinc was decanted off, most of the trifluoroacetic acid was removed in vacuo (oil pump), the remaining oil was cooled to 0xc2x0 C., and a mixture of 150 ml of 3N aqueous sodium hydroxide solution and 21 of methylene chloride, pre-cooled to 0xc2x0 C., was added, a little at a time. After insolubles were removed by filtration, the organic phase was separated off, the aqueous phase was extracted eight times using 20 ml of methylene chloride, the collected organic phases were dried over sodium sulfate, and 20 ml of 6M methanolic hydrochloric acid were subsequently added, with ice-cooling. During this process, the product precipitated in the form of the hydrochloride as a white solid, the suspension being cooled overnight to 4xc2x0 C. to bring crystallization to completion. 2.2 g of product resulted as colorless needles (50% of theory).
19 g (105.42 mmol) of 5-cyano-3,4-dimethylthiophene-2-carboxamide were suspended in 760 ml of methanol and 110 ml of 2N hydrochloric acid solution, 9.5 g of Pd on charcoal (10%) were added, and the mixture was hydrogenated at room temperature. After 4.71 of hydrogen had been taken up (4 h), methanol was distilled out in vacuo, and the aqueous phase was extracted three times with ethyl acetate and subsequently freeze-dried. 16.3 g of the required product resulted as a white solid (70.4% of theory).
a) Ethyl 5-Chloromethylisoxazole-3-carboxylate
21.2 g (210 mmol) of triethylamine were added dropwise with stirring to a mixture, cooled to 10-15xc2x0 C., of 30 g (198 mmol) of ethyl 2-chloro-2-hydroxyiminoacetate and 150 ml of propargyl chloride, stirring was continued for 1 hour at room temperature, water was subsequently added, the mixture was extracted with ether, and the organic phase was dried over magnesium sulfate and evaporated in vacuo in a rotary evaporator. The residue was distilled in vacuo at 0.5 torr, the product distilling over at 116-122xc2x0 C.
b) 5-Chloromethylisoxazole-3-carboxylic Acid
14 g (250 mmol) of potassium hydroxide were added to 47.3 g (250 mmol) of ethyl 5-chloromethylisoxazole-3-carboxylate in 150 ml of ethanol, and the reaction mixture was stirred for 6 hours at 60-70xc2x0 C. After cooling, the mixture was concentrated in vacuo, the residue was taken up in water and extracted with ether, the aqueous phase was acidified with hydrochloric acid and subsequently extracted repeatedly with ether, and the ether phase was dried over sodium sulfate and concentrated in vacuo (oil pump, 50xc2x0 C.). 31 g of the required product resulted (77% of theory)
c) 5-Chloromethylisoxazole-3-carboxylic Acid Chloride
120 g (743 mmol) of 5-chloromethylisoxazole-3-carboxylic acid together with 500 ml of thionyl chloride and 2 drops of pyridine were refluxed for 10 hours, subsequently concentrated in vacuo and then distilled at 20 torr. The product distilled at 125-1330C. 78 g resulted (58% of theory)
d) 5-Chloromethylisoxazole-3-carboxamide
Ammonia was passed for 1 hour at 10-15xc2x0 C. into a solution of 10 g (55.56 mmol) of 5-chloromethylisoxazole-3-carboxylic acid chloride in 100 ml of methylene chloride and stirring was subsequently continued at room temperature for 1 hour. After the solution had cooled to 0xc2x0 C., the precipitate was filtered off with suction and washed with a little cold methylene chloride, and the residue was extracted twice by stirring with water to remove the ammonium salts. Drying in vacuo resulted in 6.58 g of pure product as a pale powder (74% of theory)
e) 5-Aminomethylisoxazole-3-carboxamide Hydrochloride
2.44 g (15.2 mmol) of 5-chloromethylisoxazole-3-carboxamide were added to a mixture of 100 ml of concentrated ammonia solution and 72 ml of methanol, and the reaction solution was warmed to 40xc2x0 C. and constantly saturated with ammonia gas during this process. After 6 hours, the precursor was reacted. The methanol was removed in vacuo, and the aqueous phase was extracted twice using methylene chloride and subsequently evaporated to dryness in vacuo under mild conditions in a rotary evaporator. The white solid residue was employed in the coupling reactions in the form of the crude product.
2-Aminomethyloxazole-4-thiocarboxamide and 2-aminomethylthiazole-4-thiocarboxamide were prepared by the method of G. Videnov, D. Kaier, C. Kempter and G. Jung Angew. Chemie (1996) 108, 1604, where the N-Boc-protected compounds described therein were deprotected using etheric hydrochloric acid in methylene chloride.
a) Monothiooxalic Diamide
Monothiooxalic diamide was prepared starting from ethyl thiooxamidate by the method of W. Walter, K.-D. Bode Liebigs Ann. Chem. 660 (1962), 74-84.
b) 2-Carbamoyl-4-chloromethylthiazole
10 g (96 mmol) of ethyl thiooxamidate were introduced into 170 ml of n-butanol, 26 g (204 mmol) of 1,3-dichloroacetone were added, and the mixture was heated for 90 minutes at 112xc2x0 C. under nitrogen. The reaction mixture was then concentrated in vacuo and the residue was extracted by stirring with n hexane [sic] (120 ml). 10 g of pure product resulted.
c) 4-Boc-Aminomethyl-2-carbamoylthiazole
10 g (56.6 mmol) of 2-carbamoyl-4-chloromethylthiazole were introduced into an ammonia-saturated solution of 350 ml of methanol and 80 ml of 25% strength aqueous ammonia solution. The reaction mixture was warmed for 6 hours at 40-42xc2x0 C. while continuously saturating with ammonia, then concentrated in vacuo and codistilled with methanol, and the residue was subsequently extracted by stirring first with ether and then with acetone. 7.6 g of crude product which still contained a small amount of ammonium chloride were isolated. To remove this secondary product, the crude product was reacted with (Boc)2O in aqueous dioxane solution, and the protected compound was purified by means of column chromatography. This resulted in 4.95 g of pure product.
d) 4-Boc-Aminomethyl-2-cyanothiazole
4.95 g (19.24 mmol) of 4-Boc-aminomethyl-2-carbamoylthiazole were introduced into 90 ml of methylene chloride and 16.7 ml (97.44 mmol) of diisopropylethylamine, the mixture was cooled to 0xc2x0 C., a solution of 6.35 ml of trifluoroacetic anhydride in 10 ml of methylene chloride was added dropwise at 0 to 5xc2x0 C., and the mixture was subsequently warmed to room temperature (TLC check). Then, 25 ml of water were added, the mixture was stirred for 30 minutes at room temperature and brought to pH 2.5 with 10% strength citric acid solution, and the organic phase was washed repeatedly, dried using magnesium sulfate and concentrated in vacuo. 5.4 g of viscous, pale brown crude product resulted, which were employed in the next step without further purification.
e) 4-Boc-Aminomethyl-2-thiocarbambylthiazole
The crude product resulting from d) (max 19.24 mmol) was dissolved in 65 ml of pyridine and 5 ml of triethylamine, saturated with hydrogen sulfide and left to stand at room temperature over the weekend. The reaction mixture was then evaporated in vacuo in a rotary evaporator, the residue was taken up in a mixture of ether and ethyl acetate, and the mixture was washed with 10% strength citric acid solution and water, dried over magnesium sulfate and evaporated in vacuo in a rotary evaporator. 6.0 g resulted as a pale yellow solid foam.
f) 4-Aminomethyl-2-thiocarbamoylthiazole Hydrochloride
The product resulting from the above experiment was taken up in 100 ml of methylene chloride, 30 ml of approx. 5-molar etheric hydrochloric acid solution were added, and the mixture was stirred overnight at room temperature. The reaction mixture was then evaporated to dryness in vacuo in a rotary evaporator, codistilled repeatedly with ether and subsequently extracted by stirring with methylene chloride. 4.15 g of the required product resulted as a pale yellow amorphous substance.
a) xcex1-Acetylglycine Methyl Ester Hydrochloride
Potassium tert-butylate (17.8 g, 157.9 mmol) was introduced into THF (120 ml), and a solution of N-diphenylmethylideneglycine methyl ester (40 g, 157.9 mmol) in THF (60 ml) was added at xe2x88x9270xc2x0 C. After the yellowish solution had been stirred for 30 minutes at this temperature, it was added dropwise at xe2x88x9270xc2x0 C. to a solution of acetyl chloride (12.4 g, 157.9 mmol) in THF (70 ml). After the mixture had been stirred at this temperature for 1.75 hours, 3N HCl (160 ml) was added, and the yellowish suspension was stirred for a further 10 minutes at room temperature. The THF was removed at room temperature on a rotary evaporator, and the remaining aqueous phase was washed 3xc3x97 with diethyl ether. The aqueous phase was freeze-dried and the residue was extracted by stirring with methanol. The methanolic solution of the product was concentrated on a rotary evaporator at 35xc2x0 C. Yield: 26.4 g (157.9 mmol, quant., yellowish solid).
b) BOC-Gly-(xcex1-Acetyl-Gly)-OMe [sic]
BOC-Gly-OH [sic] (24.05 g, 137.27 mmol) were [sic] introduced into THF (400 ml), and triethylamine (13.87 g, 137.19 mmol) was added. The colorless solution was cooled to xe2x88x9220xc2x0 C., and a solution of isobutyl chloroformate (18.75 g, 137.28 mmol) in THF (20 ml) was added dropwise at this temperature. The colorless suspension was stirred for a further 30 minutes at xe2x88x9220xc2x0 C., and a-acetylglycine methyl ester hydrochloride (23.0 g, 137.3 mmol) was then added portionwise. After the mixture had been stirred for 30 minutes at xe2x88x9220xc2x0 C., a solution of triethylamine (13.87 g, 137.19 mmol) in THF (20 ml) was added dropwise in the course of 45 minutes. After the mixture had been stirred for 4 hours at xe2x88x9220xc2x0 C., stirring was continued for another 12 hours at RT. The residue was filtered off with suction and-washed with THF, and the combined THF phases were concentrated on a rotary evaporator.
Yield: 44.1 g (pale brown oil). 1H NMR (270 MHz, CDCl3) xcex4=1.45 (s, 9H), 2.40 (s, 3H), 3.85 (s, 3H), 3.90 (d, J=6.5 Hz, 2H), 5.25 (d, J=6.5 Hz, 1H), 7.30 (sbr, 1H).
c) Methyl 2-(N-Boc-Aminomethyl)-5-methylthiazole-4-carboxylate
BOC-Gly-(a-acetyl-Gly)-OMe [sic] (39.8 g, 138.2 mmol) was introduced into THF (400 ml), and Lawesson""s reagent (96.6 g, 238.8 mmol) was added portionwise at room temperature. The yellowish solution was then refluxed for 1.5 hours. The THF was removed on a rotary evaporator. The residue (reddish-brown oil) was extracted by stirring with diethyl ether (600 ml). The ether phase was decanted off from the undissolved brownish oil .and washed in succession with 5% strength citric acid (2xc3x97), saturated NaHCO3 solution (9xc3x97) and water (2xc3x97). After drying (MgSO4) the solvent was removed on a rotary evaporator. Yield: 22.0 g (77 mmol, 56%, brownish solid).
1H NMR (270 MHz, CDCl3) xcex4=1.50 (s, 9H), 2.75 (s, 3H), 3.95 (s, 3H), 4.55 (d, J=6.5 Hz, 2H), 5.45 (t, J=6.5 Hz, 1H). (Main rotamer relative to the Boc group).
d) 2-(N-Boc-Aminomethyl)-5-methylthiazole-4-carboxylic Acid
Methyl 2-(N-Boc-aminomethyl)-5-methylthiazole-4-carboxylate (22.0 g, 77 mmol) was dissolved in ethanol (100 ml), and a solution of LiOH (2.2 g, 92 mmol) in water (50 ml) was added. After the mixture had been stirred for 30 minutes at room temperature, the ethanol was removed on a rotary evaporator and the solution which remained was diluted with water (70 ml). The aqueous phase was washed with ethyl acetate (3xc3x97) and brought to pH 2 with 20% strength NaHSO4 solution, during which process a pale brown oil separated out. The aqueous phase was extracted with dichloromethane and the combined organic extracts were dried (MgSO4) and concentrated in vacuo. The pale brown residue was extracted by stirring in diisopropyl ether. The colorless precipitate which remained was filtered off with suction and washed with diisopropyl ether. Yield: 6.9 g (25.4 mmol, 33%, colorless solid).
1H NMR (270 MHz, DMSO-d6) xcex4=1.40 (s, 9H), 2.65 (s, 3H), 4.30 (d, J=6.5 Hz, 2H), 7.80 (t, J=6.5 Hz, 1H).
e) 2-(N-Boc-Aminomethyl)-5-methylthiazole-4-carboxamide
2-(N-Boc-Aminomethyl)-5-methylthiazole-4-carboxylic acid (6.8 g, 25 mmol) was dissolved in THF (100 ml), and triethylamine (2.53 g, 25 mmol) was added. After the mixture had been cooled to xe2x88x9220xc2x0 C., a solution of isobutyl chloroformate (3.41 g, 25 mmol) in THF (10 ml) was added dropwise. After the mixture had been stirred for 30 minutes at xe2x88x9220xc2x0 C., ammonia gas was passed into the pale brown suspension for 45 minutes. The mixture was then warmed to room temperature. The residue was filtered off with suction and extracted with THF, and the filtrates were concentrated.
Yield: 6.9 g (25 mmol, quant.). 1H NMR (270 MHz, DMSO-d6) xcex4=1.40 (s, 9H), 2.65 (s, 3H), 4.30 (m, 2H), 7.40 (sbr, 1H), 7.50 (sbr, 1H), 7.80 (t, J=6.5 Hz, 1H).
f) 4-Cyano-2-(N-Boc-aminomethyl)-5-methylthiazole
2-(N-Boc-Aminomethyl)-5-methylthiazole-4-carboxamide (6.8 g, 25 mmol) was introduced into dichloromethane (120 ml). After the mixture had cooled to 0xc2x0 C. diisopropylethylamine (15.84 g, 122.8 mmol) was added dropwise. Then, a solution of trifluoroacetic anhydride (8.25 g, 39.3 mmol) in dichloromethane (20 ml) was added dropwise at xe2x88x925xc2x0 C. in the course of 30 minutes. After the mixture had been stirred for 30 minutes at 0xc2x0 C., it was warmed to room temperature, and stirring was continued for another 12 hours. The mixture was diluted with dichloromethane (100 ml) and washed with 20% strength citric acid, saturated NaHCO3 [sic] solution and saturated NaCl solution. The organic phase was dried (MgSO4) and concentrated in vacuo. Yield: 6.3 g (25 mmol, quant.).
g) 4-Amidino-2-(N-Boc-aminomethyl)-5-methylthiazolexc3x97CH3COOH
4-Cyano-2-(N-Boc-aminomethyl)-5-methylthiazole (5.5 g, 21.74 mmol) was dissolved in methanol (15-ml), and N-acetylcysteine (4.1 g, 25.12 mmol) was added. The mixture was then warmed to 60xc2x0 C., and ammonia was passed in for 22 hours. The batch was diluted with methanol and passed over an acetate ion exchanger. The methanol was removed on a rotary evaporator and the residue extracted by stirring with acetone. The colorless residue was filtered off with suction and dried in vacuo. Yield: 4.75 g (14.4 mmol, 66%, colorless solid).
1H NMR (400 MHz, DMSO-d6)xcex4=1.40 (s, 9H), 1.80 (s, 3H), 2.60 (s, 3H), 4.35 (d, J=6.5 Hz, 2H), 7.90 (t, J=6.5 Hz, 1H).
a) N-BOC-Glycine [sic] Thioamide
N-BOC-Glycinonitrile [sic] (12.0 g, 76.8 mmol) and diethylamine (0.16 ml, 2.1 mmol) were dissolved in toluene (100 ml). The solution was cooled to xe2x88x9210xc2x0 C., saturated with hydrogen sulfide and subsequently stirred overnight at room temperature. The precipitate formed was filtered off with suction and washed with toluene. The product was dried in vacuo at 45xc2x0 C. Yield: 13.2 g (69.4 mmol, 90.3%, yellowish solid).
1H NMR (270 MHz, DMSO-d6) xcex4=1.40 (s, 9H), 3.80 (d, J=7 Hz, 2H), 7.05 (t, J=7 Hz, 1H), 9.0 (sbr, 1H), 9.65 (sbr, 1H).
b) Methyl 2-(N-BOC-Aminomethyl)-4-methylthiazole-5-carboxylate [sic]
N-BOC-Glycine [sic] thioamide (10.0 g, 52.6 mmol) was introduced into methanol (70 ml), and methyl 2-chloroacetoacetate (7.9 g, 52.6 mmol) was added. The mixture was warmed for 2 hours at 60xc2x0 C. and subsequently stirred for 48 hours at room temperature. The methanol was removed on a rotary evaporator and the residue was extracted by stirring with acetone/diethyl ether. The precipitate which remained was filtered off with suction and the filtrate was concentrated. The solid obtained from the filtrate constituted the product (pure after TLC and HPLC). Yield: 8.7 g (30.4 mmol, 57.8%). ESI-MS: 287 (M+H+).
2-(N-BOC-Aminomethyl)-4-methylthiazole-5-carboxylic [sic] Acid
Methyl 2-(N-BOC-aminomethyl)-4-methylthiazole-5-carboxylate [sic] (2.8 g, 9.74 mmol) was dissolved in 1,4-dioxane (30 ml), and 1N sodium hydroxide solution (19 ml) was added. After the mixture had been stirred for 4 hours at room temperature, the 1,4-dioxane was removed on a rotary evaporator. It was diluted with water and washed with ethyl acetate. The aqueous phase was acidified with 20% strength potassium hydrogen sulfate solution, and the precipitate obtained during this process was filtered off with suction and washed with water. The product thus obtained was dried in a vacuum drying oven at 40xc2x0 C. Yield: 2.5 g.
d) 2-(N-BOC-Aminomethyl)-4-methylthiazole-5-carboxamide [sic]
2-(N-BOC-Aminomethyl)-4-methylthiazole-5-carboxylic [sic] acid (12.6 g, 46.27 mmol) was dissolved in dichloromethane (460 ml) and dimethylformamide (0.4 ml). After the mixture had cooled to 0xc2x0 C., a solution of oxalyl chloride (6.46 g, 50.90 mmol) in dichloromethane (40 ml) was added dropwise in the course of 30 minutes. After the mixture had been stirred for 2 hours at 0xc2x0 C., it was cooled to xe2x88x9220xc2x0 C., and ammonia was passed in at this temperature until the reaction was complete. The mixture was subsequently warmed to room temperature and washed with water. The precipitate formed during this process was filtered off with suction. The organic phase was washed with 5% strength citric acid solution, dried (MgSO4) and concentrated on a rotary evaporator. The resulting solid was combined with the precipitate which had previously been filtered off and dried at 50xc2x0 C. in a vacuum drying oven. Yield: 9.8 g (36.12 mmol, 78%).
e) 2-(N-BOC-Aminomethyl)-5-cyano-4-methylthiazole [sic]
2-(N-BOC-Aminomethyl)-4-methylthiazole-5-carboxamide [sic] (11.13 g, 41.02 mmol) was suspended in dichloromethane (75 ml) and cooled to 0xc2x0 C. At this temperature, ethyldiisopropylamine (17.86 ml, 102.55 mmol) was first added, and then, slowly, a solution of trifluoroacetic anhydride (6.56 ml, 47.17 mmol) in dichloromethane (20 ml). After stirring for 1 hour, the mixture was diluted with dichloromethane and washed with 5% strength citric acid solution. After drying (MgSO4), the solvent was removed on a rotary evaporator and the crude product was purified by flash chromatography.
Yield: 6.5 g (25.66 mmol, 63%).
f) 2-(N-BOC-Aminomethyl)-4-methylthiazole-5-thioamide [sic]
2-(N-BOC-Aminomethyl)-5-cyano-4-methylthiazole [sic] (7.5 g, 29.61 mmol) was dissolved in pyridine (30 ml), and triethylamine (27 ml) was added. The solution was saturated with hydrogen sulfide at 0xc2x0 C. and then left to stand for 48 hours at room temperature. The solvent was subsequently removed on a rotary evaporator, and the residue was taken up in ethyl acetate, washed with 20% strength potassium hydrogen sulfate solution and dried over magnesium sulfate. The solvent was removed on a rotary evaporator, and the crude product was dissolved in dichloromethane and precipitated with petroleum ether. The product which had precipitated was filtered off with suction and dried in a vacuum drying oven [lacuna] 40xc2x0 C. Yield: 7.1 g (24.7 mmol, 83%).
g) 5-Amidino-2-(N-BOC-aminomethyl)-4-methylthiazole [sic]xc3x97HOAc
2-(N-BOC-Aminomethyl)-4-methylthiazole-5-thioamide [sic] (7.1 g, 24.70 mmol) was dissolved in dichloromethane (40 ml), and iodomethane (17.5 g, 123.52 mmol) was added. After the mixture had been stirred for 56 hours at room temperature, the solvent was removed on a rotary evaporator. The residue was dissolved in 10% strength methanolic ammonium acetate solution (29 ml) and stirred at 40xc2x0 C. until the reaction was complete. The solvent was removed on a rotary evaporator, the residue was extracted by stirring with dichloromethane, and the resulting solid was filtered off with suction and washed with dichloromethane. The residue was dissolved in methanol and converted into the corresponding acetate by means of an acetate-loaded ion exchanger. The solvent was removed on a rotary evaporator and the resulting reddish-brown oil was extracted by stirring with dichloromethane. During this process, the product was obtained as colorless solid which was dried in vacuo at 400C. Yield: 5.3 g (16.04 mmol, 65%).
h) 5-Amidino-2-aminomethyl-4-methylthiazolexc3x972 HCl
5-Amidino-2-(N-BOC-aminomethyl)-4-methylthiazole [sic]xc3x97HOAC (1.6 g, 4.84 mmol) was suspended in dichloromethane (20 ml), and 4M hydrochloric acid in 1,4-dioxane (4.84 ml, 19.37 mmol) was added at room temperature and the mixture was stirred for 3 hours at this temperature. The product was filtered off, washed with dichloromethane and dried in vacuo at 40xc2x0 C.
Yield: 0.73 g (3.00 mmol, 62%).
a) Ethyl 2-(N-BOC-Aminomethyl)-4-trifluoromethylthiazole-5-carboxylate [sic]
N-BOC-Glycine [sic] thioamide (5.0 g, 26.28 mmol) was dissolved in acetonitrile (60 ml), and a solution of ethyl 2-chloro-4,4,4-trifluoroacetoacetate (6.38 g, 26.28 mmol) was added dropwise at 5-10xc2x0 C. Then, the mixture was stirred for a further 30 minutes at 5xc2x0 C. and for 12 hours at room temperature. The batch was then cooled to 0xc2x0 C., and triethylamine (12 ml, 86.77 mmol) was added dropwise. After the mixture had been stirred for 20 minutes at 0xc2x0 C., the yellow suspension had changed into a clear reddish-brown solution. Then, thionyl chloride (2.1 ml, 28.89 mmol) was slowly added dropwise at 0xc2x0 C. After the mixture had been stirred for 20 minutes at 0xc2x0 C., it was warmed to room temperature for a further hour. The solvent was removed on a rotary evaporator, and the residue was taken up in water (100 ml) and extracted repeatedly with ethyl acetate. The combined organic phases were dried (Na2AO4) and concentrated. The crude product was purified by chromatography (silica gel MeOH:DCM=2:98). Yield: 2.2 g (6.4 mmol, 24.5%).
1H NMR (270 MHz, DMSO-d6) xcex4=1.30 (t, J=6.5 Hz, 3H), 1.45 (s, 9H), 4.35 (q, J=6.5 Hz, 2H), 4.45 (d, J=6.5 Hz, 2H), 7.95 (t, J=6.5 Hz, 1H).
b) 2-(N-BOC-Aminomethyl)-4-tri-fluoromethylthiazole-5-carboxamide [sic]
Ethyl 2-(N-BOC-aminomethyl)-4-trifluoromethylthiazole-5-carboxylate [sic] (15 g, 42.33 mol) was dissolved in methanol. Ammonia was passed into the solution at room temperature until all of the ester had been converted into the carboxamide. The solvent was removed on a rotary evaporator and the crude product was purified by flash chromatography. Yield: 4.6 g (14.14 mmol, 33%).
c) 2-(N-BOC-Aminomethyl)-5-cyano-4-trifluoromethylthiazole [sic]
2-(N-BOC-Aminomethyl)-4-trifluoromethylthiazole-5-carboxamide [sic] (4.6 g, 14.14 mmol) was dissolved in dichloromethane (30 ml) and cooled to xe2x88x925xc2x0 C. Ethyldiisopropylamine (4.6 g, 35.35 mmol) and a solution of trifluoroacetic anhydride (3.4 g, 16.26 mmol) in dichloromethane (10 ml) were added at this temperature. Then, the mixture was stirred for a further 2 hours at 0xc2x0 C. It was washed in succession with saturated sodium hydrogen carbonate solution and 5% strength citric acid solution. After drying (MgSO4), the solvent was removed on a rotary evaporator. The crude product was extracted by stirring with diethyl ether/petroleum ether. The supernatant was separated from the oil and concentrated on a rotary evaporator. Yield: 1.9 g (6.18 mmol, 44%).
d) 2-(N-BOC-Aminomethyl)-4-trifluoromethylthiazole-5-thioamide [sic]
2-(N-BOC-Aminomethyl)-5-cyano-4-trifluoromethylthiazole [sic] (4.6 g, 14.97 mmol) was dissolved in pyridine (20 ml), triethylamine (24 ml) was added, and the solution was saturated with hydrogen sulfide. After two days at room temperature, the solvent was removed on a rotary evaporator. The crude product was taken up in ethyl acetate and washed in succession with 20% strength sodium hydrogen sulfate solution and water. After drying (MgSO4), the solvent was removed on a rotary evaporator. The crude product was purified by flash chromatography. Yield: 2.5 g (7.32 mmol, 49%).
e) 5-Amidino-2-(N-BOC-aminomethyl)-4-trifluoromethylthiazole [sic]
2-(N-BOC-Aminomethyl)-4-trifluoromethylthiazole-5-thioamide [sic] (2.5 g, 7.32 mmol) was dissolved in dichloromethane (10 ml), and iodomethane (10.4 g, 73.24 mmol) was added. Then, the mixture was stirred for 48 hours at room temperature. After the solvent had been removed on a rotary evaporator, the residue was taken up in methanol (5 ml), and 10% strength methanolic ammonium acetate solution (8.5 ml, 10.98 mmol) was added. After the mixture had been stirred for 4 days at room temperature, the solution of the crude product was passed over an acetate-loaded ion exchanger and the solvent was removed on a rotary evaporator. The crude product was purified by flash chromatography. Yield: 0.8 g (2.08 mmol, 28%).
f) 5-Amidino-2-aminomethyl-4-trifluoromethylthiazolexc3x972 HCl
5-Amidino-2-(N-BOC-aminomethyl)-4-trifluoromethylthiazole [sic] (0.8 g, 2.08 mmol) was dissolved in dichloromethane, and a 4M solution of hydrochloric acid in 1,4-dioxane (2.1 ml, 4.2 mmol) was added. After the mixture had been stirred for 1 hour at room temperature, the solvent was removed on a rotary evaporator. The crude product obtained in this way was employed in the following reactions without further purification. Yield: 0.6 g (2.0 mmol, 97%). ESI-MS: 225 (M+H+).
a) 5-Formyl-3-methylthiophene-2-carbonitrile
112 ml (179 mmol) of a 1.6-molar solution of n-butyllithium in n-hexane were added in the course of 20 minutes to a solution of 25.1 ml (179 mmol) of diisopropylamine in 400 ml of tetrahydrofuran cooled to xe2x88x9278xc2x0 C. The solution was allowed to come to xe2x88x9235xc2x0 C., then cooled again to xe2x88x9278xc2x0 C., and a solution of 20.0 g (162 mmol) of 2-cyano-3-methylthiophene in 80 ml of tetrahydrofuran was slowly added dropwise at this temperature. During this process, the color of the solution changed to dark red. Stirring was continued for 45 minutes, 63 ml (811 mmol) of dimethylformamide were slowly added dropwise, and the mixture was stirred for another 30 minutes. For work-up, a solution of 27 g of citric acid in 160 ml of water was added at xe2x88x9270xc2x0 C. The mixture was concentrated on a rotary evaporator, 540 ml of saturated sodium chloride solution were added, and the batch was extracted three times using in each case 250 ml of diethyl ether. The combined organic extracts were dried over magnesium sulfate. After the desiccant had been filtered off, the solvent was distilled off under a water pump vacuum and the residue was purified by column chromatography (eluant hexane/ethyl acetate 4/1). This gave 23 g (94%) of the title compound. 1H NMR (270 MHz, DMSO-d6): xcex4=2.4 (s, 3H), 8.0 (s, 1H), 9.8 (s, 1H).
b) 5-Hydroxymethyl-3-methylthiophene-2-carbonitrile
5.75 g (152 mmol) of sodium borohydride were added portionwise at room temperature to a solution of 23 g (152 mmol) of 5-formyl-3-methylthiophene-2-carbonitrile in 300 ml of absolute ethanol. The reaction mixture was stirred for 5 minutes, concentrated under a water pump vacuum, taken up in ethyl acetate and extracted with 5% strength citric acid solution and with saturated sodium chloride solution, the organic phase was dried over magnesium sulfate, the desiccant was filtered off, and the solvent was distilled off at room temperature and under a water pump vacuum. This gave 24 g of the title compound as a dark red oil which still contained solvent and which was employed in the following reactions without further purification. 1H NMR (270 MHz, DMSO-d6): xcex4=2.4 (s, 3H), 4.7 (m, 2H), 5.9 (m, 1H), 7.0 (s, 1H).
c) 5-Bromomethyl-3-methylthiophene-2-carbonitrile
44 g (167 mmol) of triphenylphosphine were added to a solution of 24 g (152 mmol) of 5-hydroxymethyl-3-methylthiophene-2-carbonitrile in 180 ml of tetrahydrofuran. Then, a solution of 55 g (167 mmol) of tetrabromomethane in 100 ml of tetrahydrofuran was added. The mixture was stirred for 90 minutes at room temperature. The reaction mixture was concentrated on a rotary evaporator under a water pump vacuum, and the residue was purified by column chromatography (eluant hexane:ethyl acetate 8:2). This gave 34 g of the title compound which still contained a small amount of solvent. 1H NMR (270 MHz, DMSO-d6): xcex4=2.4 (s, 3H), 5.0 (s, 2H), 7.3 (s, 1H).
d) 5-N,N-bis(tert-Butoxycarbonyl)aminomethyl-3-methylthiophene-2-carbonitrile
5.0 g (167 mmol) of sodium hydride (80% suspension in mineral oil) was added portionwise to a solution of 33.8 g (152 mmol) of 5-bromomethyl-3-methylthiophene-2-carbonitrile in 255 ml of tetrahydrofuran, cooled to 0xc2x0 C. Then, a solution of 36.4 g (167 mmol) of di-tert-butyl iminodicarboxylate in 255 ml of tetrahydrofuran was added dropwise, during this process the temperature did not rise above 5xc2x0 C. The mixture was allowed to come to room temperature and was stirred overnight. To complete the reaction, the mixture was warmed for a further 3 hours at 35xc2x0 C. and was then left to cool to room temperature, and 510 ml of a saturated ammonium chloride solution was added slowly. The solvent was distilled off under a water pump vacuum, the residue was extracted repeatedly with ethyl acetate, and the combined organic phases were washed with saturated sodium chloride solution, dried over magnesium sulfate and concentrated on a rotary evaporator. This gave 57.6 g of an oily residue which still contained di-tert-butyl iminodicarboxylate and which was employed in the following reaction as a crude product. 1H NMR (270 MHz, DMSO-d6): xcex4=1.45 (s, 18H), 2.35 (s, 3H), 4.85 (s, 2H), 7.05 (s, 1H).
e) 5-Aminomethyl-3-methylthiophene-2-carbonitrile Hydrochloride
52.6 g of 5-N,N-bis(tert-butoxycarbonyl)aminomethyl-3-methylthiophene-2-carbonitrile (crude product of d), not more than 139 mmol) were dissolved in 950 ml of ethyl acetate and the solution was cooled to 0xc2x0 C. It was saturated with hydrogen chloride gas, during which process a white precipitate separated out after 10 minutes. The mixture was stirred for two hours at room temperature and for one hour at 30xc2x0 C., the resulting suspension was subsequently concentrated on a rotary evaporator, the residue was extracted by stirring with diethyl ether, the solvent was filtered off, and the solid residue was dried at room temperature in vacuo. This gave 24.7 g (94%) of the title compound as white powder.
1H NMR (270 MHz, DMSO-d6): xcex4=2.4 (s, 3H), 4.25 (s, 2H), 7.3 (s, 1H), 8.8-9.0 (bs, 3H). 13CNMR (DMSO-d6): 15.0 (CH3), 36.4 (CH2), 104.8 (C-2), 113.8 (CN), 131.5 (C-4), 142.8 (C-5), 149.6 (C-3).
This compound was synthesized as described for 5-aminomethyl-3-methylthiophene-2-carbonitrile, the 3-chloro-2-cyanothiophene employed having been prepared by dehydrating 3-chlorothiophene-2-carboxamide with trifluoroacetic anhydride.
a) Ethyl 2-Amino-3-cyano-4-methylthiophene-5-carboxylate
Ethyl 2-amino-3-cyano-4-methylthiophene-5-carboxylate was synthesized as described in xe2x80x9cOrganikumxe2x80x9d [Organic Chemistry], 19th Edition, Dt. Verlag der Wissenschaften, Leipzig, Heidelberg, Berlin, 1993, Chapter 6, pp.374-375, starting from 130 g (1.0 mol) of ethyl acetoacetate, 66 g (1.0 mol) of malononitrile, 32 g (1.0 mol) of sulfur and 80 g (0.92 mol) of morpholin. 1H NMR (270 MHz, DMSO-d6): xcex4=1.25 (t, 3H), 2.3 (s, 3H), 4.2 (q, 2H), 7.9 (bs, 2H).
b) Ethyl 4-Cyano-3-methylthiophene-2-carboxylate
A solution of 20.5 g (97.5 mmol) of ethyl 2-amino-3-cyano-4-methylthiophene-5-carboxylate in 600 ml of a 1:1 mixture of acetonitrile and dimethylformamide was cooled to 5xc2x0 C., and 15.7 g (146 mmol) of tert-butyl nitrite were added dropwise, during which process the temperature of the reaction mixture rose and gas was evolved vigorously. The mixture was stirred for seven hours at room temperature and concentrated on a rotary evaporator under a high vacuum, the residue was purified by column chromatography (eluant dichloromethane), and 9.1 g (48%) of the desired compound were obtained as yellow oil. 1H NMR (270 MHz, DMSO-d6): xcex4=1.3 (t, 3H), 2.55 (s, 3H), 4.3 (q, 2H), 8.8 (s, 1H).
c) 5-Hydroxymethyl-4-methylthiophene-3-carbonitrile
2.44 g (64 mmol) of lithium aluminum hydride were added portionwise at 0xc2x0 C. to a solution of 25.1 g (129 mmol) of ethyl 3-cyano-4-methylthiophene-5-carboxylate in 400 ml of tetrahydrofuran. The mixture was stirred for 5 hours at room temperature, excess reducing agent was destroyed by adding 0.5 N hydrochloric acid, and the reaction mixture was concentrated under a water pump vacuum, diluted with water and extracted three times with ethyl acetate. The combined organic phases were then washed in each case once with 0.5 N hydrochloric acid and saturated sodium chloride solution. The organic phase was dried over magnesium sulfate, the desiccant was filtered off, and the solvent was distilled off under a water pump vacuum at room temperature. The residue was purified by column chromatography (eluant dichloromethane/methanol 95:5), and 16.1 g (83%) of the desired compound were obtained as a slightly yellow oil.
1H NMR (270 MHz, DMSO-d6): xcex4=2.2 (s, 3H), 4.6 (d, 2H), 5.7 (m, 1H), 8.35 (s, 1H).
d) 5-Bromomethyl-4-methylthiophene-3-carbonitrile
30 g (115 mmol) of triphenylphosphine were added at 5xc2x0 C. to a solution of 16 g (104 mmol) of 5-hydroxymethyl-4-methylthiophene-3-carbonitrile in 300 ml of tetrahydrofuran. Then, a solution of 38 g (115 mmol) of tetrabromomethane in 100 ml of tetrahydrofuran was added. The mixture was stirred overnight at room temperature. The reaction mixture was concentrated on a rotary evaporator under a water pump vacuum and the residue was purified by column chromatography (eluant petroleum ether:dichloromethane 1:1). This gave 17 g (76%) of the title compound as yellow oil. 1H NMR (270 MHz, DMSO-d6): xcex4=2.25 (s, 3H), 5.0 (s, 2H), 8.5 (s, 1H).
e) 5-N,N-bis(tert-Butoxycarbonyl)aminomethyl-4-methylthiophene-3-carbonitrile
3.5 g (103 mmol) of sodium hydride (oil-free) was added portionwise to a solution of 17.2 g (79.5 mmol) of 5-bromomethyl-4-methylthiophene-3-carbonitrile in 250 ml of tetrahydrofuran, cooled to 0xc2x0 C. Then, a solution of 22.5 g (103 mmol) of di-tert-butyl iminodicarboxylate in 100 ml of tetrahydrofuran was added dropwise, during which process the temperature did not rise above 5xc2x0 C. The mixture was allowed to warm to room temperature and was stirred for 2 hours. 400 ml of a saturated ammonium chloride solution was added slowly. The solvent was distilled off under a water pump vacuum, and the residue was diluted with a little water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated ammonium chloride solution and with saturated sodium chloride solution, dried over magnesium sulfate and concentrated on a rotary evaporator. This gave 28 g of an oil which still contained di-tert-butyl iminodicarboxylate and was employed in the following reaction as crude product. 1H NMR (270 MHz, DMSO-d6): xcex4=1.4 (s, 9H), 1.45 (s, 9H), 2.3 (s, 3H), 4.8 (s, 2H), 8.4 (s, 1H).
f) 5-N,N-bis(tert-Butoxycarbonyl)aminomethyl-4-methylthiophene-3-thiocarboxamide
The crude product obtained in e) (not more than 79 mmol) was dissolved in 280 ml of pyridine and 140 ml of triethylamine and the solution was saturated with hydrogen sulfide at room temperature. The color of the solution, which was yellow at the beginning, changed to green. The mixture was stirred overnight at room temperature. To complete the reaction, hydrogen sulfide was passed in for a further 15 minutes and stirring was continued for two hours at room temperature. Excess hydrogen sulfide was expelled with the aid of a nitrogen stream using a washing tower. Then, the reaction mixture was concentrated on a rotary evaporator, and the concentrate was taken up in ethyl acetate, washed repeatedly with 20% strength sodium hydrogen sulfate solution, dried over magnesium sulfate and concentrated on a rotary evaporator. This gave 27 g of a pale yellow solid foam which wa s employed in the following reaction without further purification. 1H NMR (270 MHz, DMSO-d6): xcex4=1.4 (s, 18H), 2.15 (s, 3H), 4.8 (s, 2H), 7.5 (s, 1H), 9.3 (bs, 1H), 9.75 (bs, 1H).
g) 5-Aminomethyl-4-methylthiophene-3-thiocarboxamide Hydrochloride
27 g of 5-N,N-bis(tert-butoxycarbonyl)-aminomethyl-4-methylthiophene-3-thiocarboxamide (crude product of f), not more than 70 mmol) were dissolved in 400 ml of ethyl acetate and the solution was cooled to 0xc2x0 C. It was saturated with is hydrogen chloride gas, during which process a white precipitate separated out after 10 minutes. The mixture was stirred for two hours at room temperature, the precipitate was filtered off and washed with ethyl acetate, and the solid residue was dried in vacuo at room temperature. This gave 13.6 g (87%) of the title compound as white powder. EI-MS: M+=186.
a) 5-Formyl-4-chlorothiophene-3-carbonitrile
35 g (325 mmol) of tert-butyl nitrite were added dropwise at room temperature to a solution of 53.0 g (250 mmol) of 2-amino-4-chloro-5-formylthiophene-3-carbonitrile (the synthesis of this compound is described in Patent DB 3738910) in 600 ml of a 1:1 mixture of acetonitrile and dimethylformamide, during which process the temperature of the reaction mixture rose from 20xc2x0 C. to 37xc2x0 C. and vigorous evolution of gas began. The mixture was cooled to 25xc2x0 C. and stirred for 7 hours at room temperature, the black solution was concentrated on a rotary evaporator under a high vacuum, the residue was purified by column chromatography (eluant dichloromethane), and 29 g (68%) of the desired compound were obtained as yellow oil. 1H NMR (270 MHz, DMSO-d6): xcex4=9.1 (s, 1H), 10.0 (s, 1H).
b) 5-Hydroxymethyl-4-chlorothiophene-3-carbonitrile
6.3 g (166 mmol) of sodium borohydride were added portionwise at 5xc2x0 C. to a solution of 28.5 g (166 mmol) of 5-formyl-4-chlorothiophene-3-carbonitrile in 400 ml of rose slightly and the color changed to dark red. A vigorous evolution of gas was observed. After 10 minutes, the reaction mixture was concentrated under a water pump vacuum, taken up in 200 ml of ethyl acetate, extracted with 200 ml of 1 M hydrochloric acid and washed twice with in each case 250 ml of water and with saturated sodium chloride solution, the organic phase was dried over magnesium sulfate, the desiccant was filtered off, and the solvent was distilled off under a water pump vacuum at room temperature. This gave 22 g (76%) of the title compound as dark red oil which was employed in the following reactions without further purification. 1H NMR (270 MHz, DMSO-d6): xcex4=4.65 (bs, 1H), 5.95 (t, 2H), 8.6 (s, 1H).
c) 5-Bromomethyl-4-chlorothiophene-3-carbonitrile
36.1 g (137 mmol) of triphenylphosphine were added at 5xc2x0 C. to a solution of 21.7 g (125 mmol) of 5-hydroxymethyl-4-chlorothiophene-3-carbonitrile in 250 ml of tetrahydrofuran. Then, a solution of 45.6 g (137 mmol) of tetrabromomethane in 100 ml of tetrahydrofuran was added. The mixture was stirred overnight at room temperature. The precipitate was filtered off, the filtrate was concentrated on a rotary evaporator under a water pump vacuum, and the residue was purified by column chromatography (eluant petroleum ether:dichloromethane 1:1). This gave 26.0 g (88%) of the title compound as an oil.
1H NMR (270 MHz, DMSO-d6): xcex4=4.95 (s, 2H), 8.8 (s, 1H).
d) 5-N,N-bis(tert-Butoxycarbonyl)aminomethyl-4-chlorothiophene-3-carbonitrile
6.9 g (159 mmol) of sodium hydride (oil-free) was added portionwise to a solution of 25.0 g (106 mmol) of 5-bromomethyl-4-chlorothiophene-3-carbonitrile in 300 ml of tetrahydrofuran, cooled to 0xc2x0 C. Then, a solution of 34.4 g (159 mmol) of di-tert-butyl-iminodicarboxylate in 100 ml of tetrahydrofuran was added dropwise, during which process the temperature did not rise above 5xc2x0 C. The mixture was allowed to warm to room temperature and was stirred for two hours. 300 ml of a saturated ammonium chloride solution were added slowly. The solvent was distilled off under a water pump vacuum, and the residue was diluted with a little water and extracted three times with ethyl acetate. The combined organic phases were washed with saturated ammonium chloride solution and with saturated sodium chloride solution, dried over magnesium sulfate and concentrated on a rotary evaporator. This gave 51.3 g of an oil which still contained di-tert-butyl iminodicarboxylate and solvent residues and which was employed in the following reaction as crude product. 1H NMR (270 MHz, DMSO-d6): xcex4=1.4 (s, 9H), 1.45 (s, 9H), 4.8 (s, 2H), 8.65 (s, 1H).
e) 5-N,N-bis(tert-Butoxycarbonyl)aminomethyl-4-methylthiophene-3-thiocarboxamide
Some of the crude product obtained in d) (39.4 g, not more than 106 mmol) was dissolved in 400 ml of pyridine and 40 ml of triethylamine and the solution was saturated with hydrogen sulfide at room temperature. The color of the solution, which was yellow at the beginning, changed to green. The mixture was stirred overnight at room temperature. Excess hydrogen sulfide was expelled with the aid of a stream of nitrogen using a washing tower. The reaction mixture was then poured into ice-cooled 20% strength sodium hydrogen sulfate solution and extracted three times with ethyl acetate. The organic phase was then washed repeatedly with 20% strength sodium hydrogen sulfate solution, dried over magnesium sulfate and concentrated on a rotary evaporator. This gave 49.0 g of a solvent-containing residue which was employed in the following reaction without further purification. 1H NMR (270 MHz, DMSO-d6): xcex4=1.4, 1.45 (s, 18H), 4.8 (s, 2H), 7.75 (s, 1H), 9.4 (bs, 1H), 10.0 (bs, 1H).
f) 5-Aminomethyl-4-chlorothiophene-3-thiocarboxamide Hydrochloride
38.0 g of the crude product of e), not more than 93 mmol, were dissolved in 400 ml of ethyl acetate and cooled to 0xc2x0 C. The solution was saturated with hydrogen chloride gas, during which process a white precipitate separated out after 10 minutes. Since the reaction was still incomplete, 200 ml of ethyl acetate were added, and the mixture was saturated again with hydrogen chloride gas and stirred overnight at room temperature. The precipitate was filtered off, washed with petroleum ether and dried in vacuo at room temperature. This gave 21.1 g of the title compound as white powder which contained ammonium chloride as contamination. EI-MS: M+=206.
a) Ethyl Aminothioxoacetate
Hydrogen sulfide was passed to saturation at 0xc2x0 C. in a solution of 29.1 g (294 mmol) of ethyl cyanoformate and 0.4 g (0.57 ml, 5.1 mmol) of diethylamine in 20 ml of benzene, during which process the solution turned orange. The mixture was stirred over the weekend at room temperature, the reaction mixture was cooled to 0xc2x0 C., and the precipitate formed (29.1 g) was filtered off and washed with cold benzene. The mother liquor was concentrated and again cooled to 0xc2x0 C. The mixture was filtered off, the residue was washed with petroleum ether, and a further 5.7 g of the title compound were obtained as pale yellowish solid (Rf=0.7, dichloromethane/methanol 9:1). Overall yield: 89%. 1H NMR (270 MHz, DMSO-d6): xcex4=1.25 (t, J=7 Hz, 3H), 4.2 (q, J=7 Hz, 2H) 9.9 (bs, 1H, NH), 10.4 (bs, 1H, NH).
b) Ethyl Methyloxamidrazonecarboxylate
A solution of 11.93 g (13.6 ml, 259 mmol) of methylhydrazine in 100 ml of ethanol was added at room temperature dropwise to a solution of 34.5 g (259 mmol) of ethyl aminothioxoacetate in 400 ml of ethanol, during which process the temperature of the reaction mixture rose slightly. The mixture was stirred for three hours at room temperature and concentrated, and the residue was employed in reaction c) without further purification.
c) Ethyl Amino[(2-tert-butoxycarbonylaminoacetyl)methyl]-hydrazonoacetate
Activation of Boc-Gly-OH and reaction with b):
37.7 g (51.7 ml, 373 mmol) of triethylamine were added at room temperature to a solution of 54.46 g (311 mmol) of Boc-glycine in 400 ml of tetrahydrofuran. The mixture was cooled to xe2x88x925xc2x0 C., and a solution of 40.47 g (35.5 ml, 311 mmol) of ethyl chloroformate in 100 ml of tetrahydrofuran was slowly added dropwise in the course of 40 minutes. The mixture was stirred for 30 minutes at xe2x88x925xc2x0 C., the resulting precipitate was filtered off and washed with a small amount of tetrahydrofuran, and the filtrate was directly reacted further by slowly adding dropwise at room temperature a solution of the residue from b) (259 mmol) in 300 ml of tetrahydrofuran. The mixture was stirred overnight and concentrated to dryness on a rotary evaporator under reduced pressure, and the residue was purified by column chromatography (silica gel, dichloromethane/methanol 95:5, Rf=0.26). This gave 15.7 g of an oil, which was taken up in diethyl ether, and the precipitate was filtered off (8.5 g, 11%). 1H NMR (270 MHz, DMSO-d6): xcex4=1.25 (t, J=7 Hz, 3H), 1.35 (s, 9H), 2.9 (s, 3H), 3.6 (d, J=5 Hz, 2H), 4.3 (q, J=7 Hz, 2H) 6.6 (t, J=5 Hz 1H), 7.3 (bs, 2H).
d) Ethyl 5-Aminomethyl-1-methyl-1H-[1,2,4]-triazole-3-carboxylate
7.0 g (23.2 mmol) of ethyl amino[(2-tert-butoxycarbonylaminoacetyl)methyl]hydrazonoacetate were suspended in 30 ml of xylene and the suspension was immersed for 10 minutes in a silicone oil bath which had been preheated to 180xc2x0 C. Then, the solvent was distilled off directly from the reaction mixture and the residue was stirred for a further 10 minutes at 180xc2x0 C. Solvent residues were removed at 50xc2x0 C. under a high vacuum, and 6.8 g ( greater than 95%) of a dark oil were obtained which oil was employed in the following reaction without further purification. A sample was filtered through silica gel and examined by NMR spectroscopy. 1H NMR (270 MHz, DMSO-d6): xcex4=1.25 (t, J=7 Hz, 3H), 1.35 (s, 9H), 3.9 (s, 3H), 4.2-4.4 (m, 4H), 7.5 (t, J=5 Hz, 1H).
e) 5-Aminomethyl-1-methyl-1H-[1,2,4]-triazole-3-carboxamide
Ammonia gas was passed for 20 minutes at xe2x88x9210xc2x0 C. into a solution of 6.8 g (not more than 23.2 mmol) of ethyl 5-aminomethyl-1-methyl-1H-[1,2,4]-triazole-3-carboxylate in 200 ml of ethanol. Stirring was continued for one hour at 0xc2x0 C. and overnight at room temperature. Since the reaction was incomplete, the procedure of passing in gas was repeated twice more (as described above) and the mixture was stirred overnight at 0xc2x0 C. The mixture was concentrated on a rotary evaporator and the residue was purified by column chromatography (dichloromethane +5-10% methanol, Rf=0.3 in dichloromethane/methanol 9:1). This gave 4.71 g as colorless oil. 1H NMR (270 MHz, DMSO-d6): xcex4=1.4 (s, 9H), 3.85 (s, 3H), 4.3 (d, J=5 Hz, 3H), 7.4 (bs, 1H), 7.6 (bs, 1H), 7.65 (bs, J=5 Hz, 1H).
f) 5-Aminomethyl-1-methyl-1H-[1,2,4]-triazole-3-carboxamide Hydrochloride
Hydrogen chloride was passed to saturation at 5xc2x0 C. to a solution of 4.7 g (not more-than 18.4 mmol) of 5-aminomethyl-1-methyl-1H-[1,2,4]-triazole-3-carboxamide in 600 ml of ethyl acetate, during which process a white precipitate formed. The mixture was stirred overnight at room temperature and concentrated on a rotary evaporator, diethyl ether was added, the mixture was concentrated and again taken up in diethyl ether, and the precipitate was filtered off and dried. This gave 3.7 g of a white solid which still contained ammonium chloride. 1H NMR (270 MHz, DMSO-d6): xcex4=3.95 (s, 3H), 4.3 (bs, 2H), 7.6 (bs, 1H), 7.75 (bs, 1H), 8.7-8.9 (m, 2H).
a) 5-N,N-bis(tert-Butoxycarbonyl)aminomethyl-3-cyanofuran
A solution, cooled to 0xc2x0 C., of 20.5 g (0.11 mol) of 5-bromomethyl-3-cyanofuran (L. M. Pevzner, V. M. Ignat""ev, B. I. Ionin, Russ. J. of Gen. Chem. 1994, 64, 2, 125-128) in 50 ml of tetrahydrofuran was added with stirring in the course of 30 minutes at 0xc2x0 C. to a suspension of 4.8 g (0.12 mol) of sodium hydride (60% dispersion in mineral oil) in 30 ml of tetrahydrofuran. A solution of 26.2 g (121 mmol) of di-tert-butyl iminodicarboxylate in 50 ml of tetrahydrofuran was subsequently added dropwise, during which process the temperature did not climb above 5xc2x0 C. The mixture was stirred for three hours at 5-10xc2x0 C., allowed to warm to room temperature and stirred overnight. 150 ml of a saturated ammonium chloride solution were slowly added. The solvent was distilled out under a water pump vacuum, the residue was extracted four times using in each case 60 ml of ethyl acetate, and the combined organic phases were washed twice using saturated sodium chloride solution, dried over magnesium sulfate and concentrated in a rotary evaporator. After drying for three hours at room temperature in vacuo (1 mm Hg), 33.2 g of a dark syrup which still contained di-tert-butyliminodicarboxylate resulted, and this was employed as crude product in the reaction below. 1H NMR (250 MHz, d6-DMSO): xcex4=1.40, 1.45 (s, 18H), 4.70 (s, 2H), 6.70 (s, 1H), 8.6 (s, 1H).
b) 5-Aminomethyl-3-cyanofuran Hydrochloride
12.89 g of 5-N,N-bis(tert-butoxycarbonyl)aminomethyl-3-cyanofuran (crude product from a) were dissolved in 80 ml of ethyl acetate and cooled to xe2x88x9210xc2x0 C. The mixture was saturated with hydrogen chloride gas, a white precipitate separating out after 15 minutes. The mixture was allowed to come to room temperature and was stirred for two hours, the resulting suspension was subsequently concentrated in a rotary evaporator, the residue (7 g) was extracted by stirring with diethyl ether, solvent was removed by filtration, and the solid residue was dried in vacuo at room temperature. 5 g (79%) of the title compound resulted as a pale ochre powder. 1H NMR (250 MHz, d6-DMSO): xcex4=4.15 (bs, 2H), 7.0 (s, 1H), 8.6-8.9 (m, 4H).
a) 5-Cyano-1-methylpyrrole-2-carbaldehyde
1-Methylpyrrole was converted into 2-cyano-1-methylpyrrole by reaction with chlorosulfonyl isocyanate and dimethylformamide in acetonitrile (see, for example, C. E. Loader et al. Can. J. Chem. (1981), 59, 2673-6).
Diisopropylamine (17.5 ml, 124.38 mmol) was introduced into THF (100 ml) under nitrogen. N-Butyllithium solution in hexane (15% strength, 75.9 ml, 124.38 mmol) was added dropwise at xe2x88x9278xc2x0 C. The mixture was subsequently stirred for 45 minutes at xe2x88x9220xc2x0 C. and then cooled again to xe2x88x9278xc2x0 C. At this temperature, a solution of 1-methylpyrrole-2-carbonitrile (12 g, 113.07 mmol) in THF (50 ml) was added dropwise. After stirring for 45 minutes at xe2x88x9278xc2x0 C., DMF (43.9 ml, 546.46 mmol) was added dropwise, and the mixture was stirred at this temperature for a further 2 hours. After addition of citric acid monohydrate (20.56 g), the mixture was warmed to room temperature, and water (112 ml) was added. The THF was removed in a rotary evaporator, and the aqueous phase was saturated with sodium chloride and extracted with diethyl ether (3xc3x97200 ml). The combined organic phases were washed with saturated sodium chloride solution and dried over sodium sulfate. The solvent was removed in a rotary evaporator and the crude product was purified by means of flash chromatography (silica gel, dichloromethane). Yield: 8.25 g (54%).
1H [sic] NMR (CDCl3) xcex4=4.1 (s, 3H), 6.8 (d, 1H), 6.9 (d, 1H), 9.7 (s, 1H).
b) 5-Hydroxymethyl-1-methylpyrrole-2-carbonitrile
The product obtained in accordance with a) (8.2 g, 61.1 mmol) was dissolved in ethanol (200 ml), and sodium borohydride (2.31 g, 61.13 mmol) was added at xe2x88x9210xc2x0 C. After stirring for 1.5 hours at 0-5xc2x0 C., the solvent was removed in a rotary evaporator, and ice-water and 20% strength sodium hydrogen sulfate solution were added to the residue. The aqueous phase was extracted with ethyl acetate. The combined organic phases were washed to neutrality with saturated sodium hydrogen carbonate solution and water and dried over sodium sulfate. The solvent was removed in a rotary evaporator and the crude product was purified by means of flash chromatography (silica gel, dichloromethane/methanol=97.5/2.5). Yield: 7.6 g (91%). 1H [sic] NMR (CDCl3) xcex4=1.9 (t, 1H), 3.75 (s, 3H), 4.6 (d, 2H), 6.1 (d, 1H), 6.7 (d, 1H).
c) 5-Azidomethyl-1-methylpyrrole-2-carbonitrile
The product obtained in accordance with b) (7.5 g, 55.08 mmol) was dissolved in DMF (220 ml), and triphenylphosphine (43.34 g, 165.25 mmol) was added at 0xc2x0 C. After stirring for 5 minutes at this temperature, tetrabromomethane (54.8 g, 165.25 mmol) was added. The mixture was subsequently stirred for 30 minutes at 0xc2x0 C. and for 1.5 hours at room temperature. After cooling to 0xc2x0 C., sodium azide (4.37 g, 67.21 mmol) was added. The mixture was subsequently stirred for 4.5 hours at room temperature. Saturated sodium chloride solution was added dropwise at 0xc2x0 C., and the batch was diluted with ethyl acetate. The organic phase was separated off, and the aqueous phase was extracted with diethyl ether. The combined organic phases were washed with water and dried over sodium sulfate. The solvent was removed in a rotary evaporator and the crude product was purified by means of flash chromatography (silica gel, ethyl acetate/hexane=1/20).
Yield: 5.6 g (63%). 1H [sic] NMR (CDCl3) xcex4=3.75 (s, 3H), 4.35 (s, 2H), 6.2 (d, 1H), 6.7 (d, 1H).
d) 5-Aminomethyl-1-methylpyrrole-2-carbonitrile
The product obtained in accordance with c) (4.71 g, 29.25 mmol) was dissolved in methanol (100 ml), and palladium on charcoal (10%, 1 g) was added. The mixture was subsequently hydrogenated with hydrogen under 1 atmosphere for 4 hours. The catalyst was removed by filtration through Celitee and the filtrate was evaporated in a rotary evaporator. The residue was extracted by stirring with dichloromethane/diethyl ether=1/1. The product was filtered off with suction and dried at 35xc2x0 C. in a vacuum drying oven.
Yield: 2.7 g (68%). 1H [sic] NMR (CDCl3) xcex4=3.75 (s, 3H), 3.85 (s, 2H), 6.05 (d, 1H), 6.7 (d, 1H).
a) 5-Cyano-1-methylpyrrole-3-carbaldehyde
Aluminum trichloride (24.24 g, 180.86 mmol) was dissolved in nitromethane/dichloromethane (1/1, 320 ml), the solution was cooled to xe2x88x9220xc2x0 C., and 1-methylpyrrole-2-carbonitrile (8 g, 75.36 mmol) was added. xcex1,xcex1-Dichlorodimethyl ether (10.4 g, 90.43 mmol), dissolved in dichloromethane (42 ml), was subsequently added dropwise. After stirring for 4 h at 0xc2x0 C., the batch was poured onto ice (200 g). The aqueous phase was extracted with diethyl ether. The combined organic phases were washed until neutral with saturated sodium hydrogen carbonate solution, water and saturated sodium chloride solution. After drying over sodium sulfate, the solvent was removed in a rotary evaporator. The crude product was employed in the reactions below without further purification.
Yield: 9.2 g (91%). 1H [sic] NMR (CDCl3) xcex4=3.8 (s, 3H); 7.2 (s, 1H); 7.4 (s, 1H); 9.85 (s, 1H).
b) Starting from 5-Cyano-1-methylpyrrole-3-carbaldehyde; 4-aminomethyl-1-methylpyrrole-2-carbonitrile was synthesized analogously to the synthesis of 5-aminomethyl-1-methylpyrrole-2-carbonitrile. However, the 4-azidomethyl-1-methylpyrrole-2-carbonitrile was advantageously reduced in a Staudinger reaction (see S. Nagarajan et al. J. Org. Chem. 1987, 52, 5044-6).
1H [sic) NMR (DMSO-d6) xcex4=3.77 (s, 3H), 3.84 (sbr, 2H), 7.00 (sbr, 1H), 7.26 (s, 1H), 8.05 (sbr, 2H).
a) 4-Cyano-1-methylpyrrole-2-carbaldehyde
1-Methylpyrrole-2-carbaldehyde (10 g, 91.6 mmol) was dissolved in acetonitrile (100 ml) and cooled to xe2x88x9245xc2x0 C. Chlorosulfonyl isocyanate (38.9 g, 274.9 mmol) in acetonitrile (40 ml) was added dropwise in the course of 40 minutes. The mixture was subsequently stirred for 12 hours at room temperature. After dropwise addition of dimethylformamide (35 ml), the mixture was warmed to 50xc2x0 C. for 1 hour. After cooling to room temperature, the reaction mixture was poured onto ice (200 ml) and 2N sodium hydroxide solution (286 ml). The precipitate formed was filtered off with suction. The filtrate was extracted with diethyl ether. The combined ether phases were washed until neutral with dilute sodium hydrogen carbonate solution and water and dried over sodium sulfate. The solvent was distilled out in a water pump vacuum and the residue was combined with the precipitate previously obtained. Recrystallization from petroleum ether gave 4-cyano-1-methylpyrrole-2-carbaldehyde (4.3 g) (see, for example, C. E. Loader et al. Can. J. Chem. (1981), 59, 2673-6) 1-H [sic] NMR (CDCl3) xcex4=4.0 (s, 3H); 7.2 (s, 1H); 7.3 (s, 1H); 9.6 (s, 1H).
13-C [sic] NMR (CDCl3) xcex4=37.4; 94.1; 114.7; 125.8; 132.2; 135.8; 179.7.
b) Starting From 4-Cyano-1-methylpyrrole-2-carbaldehyde, 5-Aminomethyl-1-methylpyrrole-3-carbonitrile was Prepared Analogously to the Synthesis of 5-Aminomethyl-1-methylpyrrole-2-carbonitrile.
1H [sic] NMR (DMSO-d6) xcex4=3.6 (s, 3H), 3.8. (s, 2H), 4.2 (sbr, 2H), 6.4 (s, 1H), 7.6 (s, 1H).
a) N-Boc-5-Aminomethyl-3-cyano-1,2,4-oxadiazole
Ethyl N-Boc-5-aminomethyl-1,2,4-oxadiazole-2-carboxylate (S. Borg et al. J. Org. Chem. 1995, 60, 3112-20) was dissolved in methanol (50 ml). Ammonia was passed into this solution at xe2x88x9210xc2x0 C. to RT until the reaction was complete. The solvent was removed in a rotary evaporator. The resulting crude product was dissolved in dichloromethane (70 ml), and diisopropylethylamine (2.9 ml, 16.55 mmol) was added at xe2x88x925xc2x0 C. Trifluoroacetic anhydride (1.06 ml, 7.61 mmol), dissolved in dichloromethane (10 ml), was subsequently added dropwise. After stirring for 1.5 hours at 0xc2x0 C., the batch was diluted with dichloromethane, washed 2xc3x97 with saturated sodium hydrogen carbonate solution, 2xc3x97 with 5% strength citric acid solution and 1xc3x97 with saturated sodium chloride solution and then dried over sodium sulfate. The solvent was removed in a rotary evaporator and the crude product was purified by chromatography (silica gel, dichloromethane:methanol=97.5:2.5). Yield: 1.2 g (80%).
b) 5-Aminomethyl-3-cyano-1,2,4-oxadiazole Hydrochloride
The product obtained in accordance with a) (0.9 g, 4.0 mmol) was dissolved in dichloromethane (45 ml), and 4M hydrochloric acid in dioxane (3.9 ml, 15.61 mmol) was added at RT. After stirring for 16 hours at RT, the solvent was removed in a rotary evaporator. Yield: 645 mg (100%).
1-H [sic] NMR (DMSO-d6) xcex4=4.6 (s, 2H), 9.2 (s, 3H).
a) Methyl 1-Methyl-5-amidopyrazole-3-carboxylate
1-Methyl-3-methoxycarbonylpyrazole-5-carboxylic acid chloride (prepared from 3.7 g, 20.09 mmol, of 1-methyl-3-methoxycarbonyl-3-carboxylic acid, J. Org. Chem. 1989, 54, 428) was dissolved in toluene and the solution was cooled to xe2x88x9210xc2x0 C. Ammonia was subsequently passed in at xe2x88x9210xc2x0 C. to 0xc2x0 C. until the reaction was complete. The solvent was removed in a rotary evaporator. The residue was taken up in ethanol. After stirring for 15 minutes, the ethanol was removed in a rotary evaporator, and the residue was dissolved in warm water and precipitated by cooling the solution to 0xc2x0 C. The precipitate was filtered off with suction, washed with acetone and dried in vacuo at 45xc2x0 C. Yield: 1.5 g (41%).
b) Methyl 1-Methyl-5-cyanopyrazole-3-carboxylate
The product obtained in accordance with a) (1.5 g, 8.19 mmol) were [sic] taken up in dichloromethane (20 ml). Diisopropylethyl- amine (3.85 ml, 22.11 mmol) was added at xe2x88x9210xc2x0 C., and a solution of trifluoroacetic anhydride (1.3 ml, 9.44 mmol) in dichloromethane (5 ml) was added dropwise at this temperature in the course of 45 minutes. Stirring was subsequently continued for 1 hour at 0xc2x0 C. The batch was diluted with dichloromethane and washed 2xc3x97 with saturated sodium hydrogen carbonate solution, 2xc3x97 with 5% strength citric acid solution and 1xc3x97 with saturated sodium chloride solution. After drying over sodium sulfate, the solvent was removed in a rotary evaporator. Yield: 1.35 g (100%).
c) 1-Methyl-5-cyanopyrazole-3-carboxamide
The product obtained in accordance with b) (1.35 g, 8.19 mmol) was introduced into methanol (50 ml) and cooled to xe2x88x9210xc2x0 C. Ammonia was subsequently passed in in the course of 8 hours. After stirring for 12 hours at room temperature, reaction of the precursor had ended. The product which had precipitated was filtered off with suction, washed with cold methanol and dried in vacuo. Yield: 1.22 g (100%).
1-H [sic] NMR (DMSO-d6) xcex4=4.0 (s, 3H), 7.4 (s, 1H), 7.5 (s, 1H), 7.8 (s, 1H).
d) 1-Methyl-5-aminomethylpyrazole-3-carboxamide
The product obtained in accordance with c) (0.4 g, 2.66 mmol) was dissolved in acetic acid (30 ml) and 10% palladium on charcoal (78 mg) was added. The mixture was subsequently hydrogenated at room temperature under atmospheric pressure until the reaction was complete. The catalyst was removed by filtration through Celite(copyright) and the solvent was removed in a rotary evaporator. Yield: 0.4 g (100%), FAB-MS (M+H+): 155.
a) Methyl 1-Methyl-3-amidopyrazole-5-carboxylate
1-Methyl-5-methoxycarbonylpyrazole-3-carbonyl chloride (synthesized from 4.17 g, 22.6 mmol, of 1-methyl-5-methoxycarbonyl-3-carboxylic acid, J. Org. Chem. 1989, 54, 428) was dissolved in toluene and the solution was cooled to xe2x88x9210xc2x0 C. Then, ammonia was passed in at xe2x88x9210xc2x0 C. to 0xc2x0 C. until the reaction was complete. The solvent was removed in a rotary evaporator. The residue was taken up in ethanol. After the mixture had been stirred for 15 minutes, the ethanol was removed in a rotary evaporator, and the residue was dissolved in warm water and precipitated by cooling to 0xc2x0 C. The precipitate was filtered off with suction, washed with acetone and dried in vacuo at 45xc2x0 C. Yield: 3.36 g (18.4 mmol, 81%).
1H NMR (270 MHz, DMSO-d6) xcex4=3.85 (s, 3H), 4.15 (s, 3H), 7.20 (s, 1H), 7.4 (sbr, 1H), 7.7 (sbr, 1H).
b) Methyl 1-Methyl-3-cyanopyrazole-5-carboxylate
The product obtained in a) (3.36 g, 18.4 mmol) was reacted similarly to the method described above for the synthesis of methyl 1-methyl-cyanopyrazole-3-carboxylate. Yield: 2.59 g (15.7 mmol, 85%).
1H NMR (250 MHz, DMSO-d6) xcex4=3.90 (s, 3H), 4.15 (s, 3H), 7.60 (s, 1H).
c) 1-Methyl-3-cyanopyrazole-5-carboxamide
The product obtained in b) (2.56 g, 15.5 mmol) was reacted similarly to the method described above for the synthesis of 1-methyl-5-cyanopyrazole-3-carboxamide. Yield: 2.3 g (15.3 mmol, 99%).
1H NMR (250 MHz, DMSO-d6) xcex4=4.15 (s, 3H), 7.45 (s, 1H), 7.70 (sbr, 1H), 8.15 (sbr, 1H).
d) 1-Methyl-3-aminomethylpyrazole-5-carboxamidexc3x97HCl
The product obtained in c) (1.0 g, 6.7 mmol) was reacted similarly to the method described above for the synthesis of 1-methyl-5-aminomethylpyrazole-3-carboxamide. Yield: 1.5 g (5.6 mmol, 83%).
1H NMR (270 MHz, DMSO-d6) xcex4=4.00 (q, J=6.5 Hz, 2H), 4.10 (s, 3H), 6.90 (s, 1H), 7.60 (sbr, 1H), 8.05 (sbr, 1H), 8.25 (sbr, 3H).
The product can be converted into the corresponding hydrochloride by repeatedly treating it with HCl in 1,4-dioxane and subsequently concentrating the mixture.